Method and device for acquiring thermodynamic and kinetic parameters of RNA (Ribonucleic Acid) pseudoknot structure

Abstract

The invention provides a method and device for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure. The method comprises the steps that a molecular structure is determined, and motion tracks of the molecular structure at different temperatures and different moments are simulated based on molecular dynamics simulation software; obtaining the RMSD and torsion angle of geometric parameter basic groups of specific basic groups in a loop region / Stem region in the molecular structure and a dihedral angle formed by the specific basic groups based on the motion trail; judging the RMSD and the torsion angle of the specific basic group at different moments and a two-state condition range satisfied by a dihedral angle formed by each specific basic group, and determining the state of the specific basic group; according to the state of the specific basic group, carrying out statistics on the average life, times and probability of the specific basic group in a closed state and an open state under the total time of simulation; thermodynamic parameters and dynamic parameters of the specific basic group are calculated according to the average service life, the number of times and the probability of the specific basic group in the closed state and the open state. The technical problem that the precision of thermodynamic parameter calculation of a loop region is not high is solved.

Description

Technical Field

[0001] The present application belongs to the technical field of molecular dynamics simulation, and relates to a method and device for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure. Background Art

[0002] RNA molecules perform a variety of biological functions in cells, from the transfer of genetic information, to serving as an intermediate in the transfer of DNA to proteins, and to enzyme catalysis. The three-dimensional structure of RNA molecules is largely determined by the interaction between base-paired nucleotide pairs. The secondary structure of an RNA molecule is a collection of specific bases in a three-dimensional structure. Therefore, the secondary structure can be used as a basis for predicting higher structural levels. The primary structural information of RNA can be obtained in large quantities through experiments, but obtaining the three-dimensional structure of RNA through experiments will make it costly and difficult, and not all three-dimensional structures can be measured experimentally.

[0003] With the continuous development of computers, people can use theoretical methods to predict the structure of RNA. The accuracy of the prediction depends on the assumptions of the folding structure and the potential energy model. In structure prediction, a common method is the maximum base pairing method, which is an algorithm developed based on the structural conservation of RNA. It mainly relies on the alignment of sequences. However, the main disadvantage of this method is the sequence alignment of conservative regions. Another method is the minimum free energy method, which is to find a complementary structure in an energy-stable state. Using this method, a lot of software has been developed. With continuous development and research, there are more and more methods for predicting RNA secondary structure.

[0004] Pseudoknot is a typical tertiary structure. Pseudoknot has different roles in biological functions, such as self-cleavage of introns, formation of catalytic cores of various ribozymes, telomerase, riboswitches and ribosomal frameshifting. The biological function of pseudoknot is directly related to its folding stability and conformational changes. In RNA pseudoknot, stability and folding thermodynamics are mainly determined by the interaction between the loop region and the stem region. For example, pseudoknot involves several intermediates in the folding process. Studying how RNA pseudoknot structure folds and forms, misfolded intermediates may be formed during the folding process, so establishing a correct model is very important for predicting the structure of RNA pseudoknot. But the biological function of pseudoknot is directly related to its folding stability and conformational changes. In RNA, pseudoknot, stability and folding thermodynamics are mainly determined by the interaction between the loop region and the stem region. Similar to previous studies on the determination and estimation of thermodynamic parameters of classical RNA secondary structures, experiments have determined the thermodynamic parameters of some model structures of RNA pseudoknots, indicating that the ion concentration in the solution is very important for the stability of RNA pseudoknots. And the conformational transitions involved in the folding process in pseudoknots may contain intermediate steps, mainly in the process of forming hairpins. Therefore, it is very difficult to study the contribution of the loop region of the pseudoknot structure to the structural stability, especially for pseudoknots containing at least two loop structures. For the pseudoknot problem, researchers have developed many algorithms over the years to try to make accurate predictions. However, due to the compact structure of RNA pseudoknots, non-bonded interactions between atoms will form in the pseudoknot, such as van der Waals interaction, electrostatic interaction, and volume repulsion. Therefore, it is inaccurate to use thermodynamic parameters in the secondary structure to predict the pseudoknot structure. Summary of the invention

[0005] The present application provides a method and device for obtaining thermodynamic and kinetic parameters of RNA pseudoknot structures, which solves the technical problems that in the prior art, only the thermodynamic parameters of some base complementary pairing can be obtained through experiments, but the thermodynamic parameters of the loop region cannot be obtained, and in the prediction of RNA pseudoknot structures, the thermodynamic parameters of the loop region calculated by different algorithms are not accurately calculated.

[0006] This application adopts the following technical solutions:

[0007] The present application provides a method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure, comprising:

[0008] Determine the molecular structure and simulate the motion trajectory of the molecular structure at different temperatures and times based on molecular dynamics simulation software. The motion trajectory includes the speed and position of the molecular structure at different times.

[0009] Based on the motion trajectory, the geometric parameters of specific bases in the loop region / Stem region in the molecular structure during the open-closed state transition are obtained. The geometric parameters include the RMSD of the base, the torsion angle, and the dihedral angle formed by each specific base.

[0010] The RMSD, torsion angle and dihedral angle range of the specific bases at different times are judged to determine the state of the specific bases.

[0011] The average lifetime, number of times and probability of a specific base in the closed state and the open state during the total time of the statistical simulation based on the state of the specific base.

[0012] The thermodynamic parameters and kinetic parameters of the specific base are calculated based on the average lifespan, number of times and probability of the specific base in the closed state and the open state, wherein the thermodynamic parameters include the free energy difference and the linear relationship between the free energy difference and the temperature; the kinetic parameters include the closing rate and the opening rate.

[0013] Optionally, the molecular structure is determined, and the motion trajectory of the molecular structure at different temperatures and at different times is simulated based on molecular dynamics simulation software, including: downloading the molecular structure from the PDB library and importing it into the molecular dynamics simulation system.

[0014] Set the periodic boundary conditions, motion equations, field forces, temperature, pressure, and running time of the molecular dynamics simulation system, run the molecular dynamics simulation system, and obtain the motion trajectory of the molecular structure at different temperatures and times.

[0015] Optionally, running a molecular dynamics simulation system to obtain the motion trajectory of the molecular structure at different temperatures and at different times includes: selecting a molecular dynamics simulation mode, setting the temperature, pressure, and total simulation time.

[0016] Given the initial position and initial velocity of a specific base, the velocity Verlet algorithm is used to solve the Newton's motion equations in the molecular dynamics simulation mode to obtain the motion trajectory of the molecular structure at different temperatures and times.

[0017] Optionally, obtaining geometric parameters of specific bases in the loop region / Stem region in the molecular structure during the open-closed state transition based on the motion trajectory includes: counting the positions of each specific base in the loop region / Stem region in the molecular structure at different times.

[0018] The RMSD, torsion angle, and dihedral angle of each specific base are calculated based on the position of each specific base.

[0019] Optionally, determining the range of two-state conditions satisfied by the RMSD, torsion angle, and dihedral angles of specific bases at different times to determine the state of the specific bases includes: if the RMSD region of a specific base constituting a base pair at any time is between The RMSD region of another specific base is ~ When the dihedral angle formed by the two specific bases ranges from ∼-80° to ∼-140°, and the torsion angle is between ∼-50° and ∼-100°, the state of the two specific bases is determined to be a closed state.

[0020] Optionally, determining the range of two-state conditions satisfied by the RMSD, torsion angle, and dihedral angles of specific bases at different times, and determining the state of the specific bases further comprises: if the RMSD of a specific base constituting a base pair at any time is greater than Another specific base has an RMSD greater than When the dihedral angle formed by the two specific bases is between -140° and 100°, and the torsion angle is between 50° and 100°, the state of the two specific bases is determined to be the open state.

[0021] Optionally, the average lifespan, number of times and probability of a specific base in a closed state and an open state during the total time of a state statistical simulation of a specific base include: separately counting the number of times each specific base appears in a closed state or an open state during the total time of each simulation at each simulation temperature.

[0022] The average life span and probability of the closed state are calculated based on the number of closed states and the time each time the closed state is maintained, or the average life span and probability of the open state are calculated based on the number of open states and the time each time the open state is maintained.

[0023] Optionally, calculating the thermodynamic parameters and kinetic parameters of a specific base based on the average lifetime, number and probability of a specific base in a closed state and an open state includes: calculating the thermodynamic parameter free energy difference of base pairs formed by the specific base at different temperatures based on the enthalpy variable and entropy variable of the transition between the closed state and the open state of the specific base.

[0024] The linear relationship between the free energy difference and temperature is calculated based on the probability of the closed and open states of a specific base;

[0025] Based on the average lifetime and number of closed / open states, the kinetic parameters closing rate / opening rate of the specific base are calculated respectively.

[0026] The present application also provides a device for acquiring thermodynamic and kinetic parameters of an RNA pseudoknot structure, including: a motion trajectory acquisition module, which is used to determine the molecular structure, and simulate the motion trajectory of the molecular structure at different temperatures and different times based on molecular dynamics simulation software, wherein the motion trajectory includes the speed and position of the molecular structure at different times.

[0027] The geometric parameter calculation module is used to obtain the geometric parameters of specific bases in the loop region / Stem region in the molecular structure during the open-closed state transition based on the motion trajectory. The geometric parameters include the RMSD of the base, the torsion angle, and the dihedral angle formed by each specific base.

[0028] The state determination module is used to determine the RMSD, torsion angle and dihedral angle range of specific bases at different times and determine the state of the specific bases.

[0029] The calculation module is used to calculate the average lifespan, number of times and probability of a specific base in a closed state and an open state during the total time of a statistical simulation of the state of the specific base.

[0030] The output module is used to calculate the thermodynamic parameters and kinetic parameters of the specific base according to the average lifespan, number and probability of the specific base in the closed state and the open state, wherein the thermodynamic parameters include the free energy difference and the linear relationship between the free energy difference and the temperature; the kinetic parameters include the closing rate and the opening rate.

[0031] The present application provides a computer device, including a memory, a processor, and a computer program stored in the memory and executable on the processor. When the processor executes the computer program, a method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure as described in any one of the above method embodiments is implemented.

[0032] The present application provides a computer-readable storage medium, which stores a computer program. When the computer program is executed by a processor, it implements a method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure in any one of the above method embodiments.

[0033] The beneficial effects of this application are:

[0034] The present application proposes a method and device for obtaining the thermodynamic and kinetic parameters of an RNA pseudoknot structure. The method uses molecular dynamics simulation to simulate the motion process of molecules at different temperatures to obtain motion trajectories; the geometric parameters of specific bases in the loop region / Stem region of the molecular structure during the open-closed state transition process are determined according to the speed and position in the motion trajectory, and the average lifespan, number of times, and probability data of the specific bases in the closed state and the open state are calculated based on the geometric parameters, and the thermodynamic parameters of the specific bases including the free energy difference, the linear relationship between the free energy difference and the temperature are calculated based on these data; the kinetic parameters include the closing rate and the opening rate, which lays the foundation for studying the influence of temperature changes on base pairs in the loop region / Stem region of the molecular structure according to the kinetic parameters and thermodynamic parameters of the specific bases, and the empirical formula is used for calculation in the process of calculating the kinetic parameters and thermodynamic parameters, so that the data acquisition is more accurate and close to the actual data. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 A step diagram of a method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure provided in this application.

[0036] Figure 2 A detailed structure diagram of HIV (PDBID: 1hvu) provided for this application.

[0037] Figure 3 A schematic diagram of an H-type pseudoknot structure provided in this application.

[0038] Figure 4 A graph showing the relationship between the RMSD of a specific base U8 provided in this application and the simulation time.

[0039] Figure 5 A schematic diagram of the dihedral angle formed by a specific base U8 and a specific base U9 provided in the present application.

[0040] Figure 6 The present application provides a simulation temperature of 360K, a total simulation time of 10 μs, and a graph showing the relationship between the RMSD of a specific base 27 and the simulation time.

[0041] Figure 7 A simulation temperature of 360K is provided for this application, and a distribution diagram of the RMSD of a specific base 27 during the entire simulation process.

[0042] Figure 8 A simulation temperature of 360K is provided for this application, and a distribution diagram of the phosphate backbone torsion angle of a specific base 27 is provided during the entire simulation process.

[0043] Fig. 9A probability diagram of the closed state during simulation for all simulated temperatures provided in this application.

[0044] Fig.10 A kind of Linear relationship graph with the inverse of temperature (1 / T).

[0045] Fig.11 A bar graph of the average lifetimes of the open and closed states at each simulation temperature is provided for this application.

[0046] Fig.12 The present application provides a lifetime distribution diagram of the open state and the closed state at a simulation temperature of 360K.

[0047] Fig.13 Schematic diagram of a device for acquiring thermodynamic and kinetic parameters of an RNA pseudoknot structure provided in this application. DETAILED DESCRIPTION

[0048] The present application is described in detail below with reference to the accompanying drawings and specific implementation methods.

[0049] The present application provides a method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure, the method comprising steps S1-S5.

[0050] S1: Determine the molecular structure and simulate the motion trajectory of the molecular structure at different temperatures and at different times based on molecular dynamics simulation software. The motion trajectory includes the speed and position of the molecular structure at different times.

[0051] Optionally, the molecular structure is determined, and the motion trajectory of the molecular structure at different temperatures and different times is simulated based on molecular dynamics simulation software, wherein the motion trajectory includes the speed and position of the molecular structure at different times, including: downloading the molecular structure from the PDB library and importing it into the molecular dynamics simulation system.

[0052] Set the periodic boundary conditions, motion equations, field forces, temperature, pressure, and running time of the molecular dynamics simulation system, run the molecular dynamics simulation system, and obtain the motion trajectory of the molecular structure at different temperatures and times.

[0053] In one embodiment, first, before performing a molecular dynamics simulation, the initial structure of the simulated molecule must be selected. The structure of the molecule can be downloaded from the PDB (protein data bank) library. Then, a suitable water box size is established. The size and shape of the box are determined by three vectors, and periodic boundary conditions are generally selected. Then, ions are randomly added to the solution to ensure the accuracy of the simulation and to ensure that the positions of the ions are randomly distributed. Finally, a molecular dynamics simulation system is selected based on the molecular dynamics simulation software to determine the appropriate field force. This application takes a crystal structure downloaded from the PDB library (PDB ID: 1hvu) as an example. Figure 2 As shown in the figure, the crystal structure of 1hvu contains two Stem regions (S1 region and S2 region) and two loop regions (L1 region and L2 region), which is the simplest H-type pseudoknot structure. The S1 region has 5 bases stacked, the S2 region contains 6 bases stacked, and the L1 region and L2 region have 2 nucleotides and 3 nucleotides respectively. L1 is in the major groove formed by the S2 region, and the L2 region passes through the minor groove formed by the S1 region.

[0054] In order to compare the thermodynamic and kinetic parameters of the 3'-AAUU-5' sequence without loop, the U26A in the S2 region was mutated. In addition, 26 Na+ ions were added to the molecular dynamics simulation system to make the entire simulation system charge neutral, and then Na+ ions and Cl- ions were added at random initial positions, and the concentration of ions in the field was 0.5M. The TIP3P water model was used and the SETTLE algorithm was used to keep the water molecules completely rigid. The simulation water cube box was set to 64*74* Periodic boundary conditions were used to ensure that there were three or four solvent layers in each direction. The final simulation system contained 26,356 atoms.

[0055] Secondly, run the molecular dynamics simulation system to obtain the motion trajectory of the molecular structure at different temperatures and times, including: selecting the molecular dynamics simulation mode, setting the temperature, pressure, and total simulation time; given the initial position and initial velocity of a specific base, using the velocity Verlet algorithm to solve the Newton's motion equations under the molecular dynamics simulation mode to obtain the motion trajectory of the molecular structure at different temperatures and times.

[0056] Specifically, the equation of motion of the molecular structure is selected in the system. According to the classical equation of motion, a simple atomic system can be written. For a system containing N interacting atoms, molecular dynamics simulation solves its Newtonian equation of motion:

[0057]

[0058] Among them, F iis the force acting on the atom, r i (i=1, 2, 3, ..., N) is the coordinate set of the entire 3N atoms.

[0059] The force on the atom is the negative of the inverse of a potential energy function, which is used to generate the potential energy and its derivative, that is, the system of equations for the force.

[0060]

[0061] Where U is the potential energy between atoms.

[0062] It should be noted that the equation of motion of the molecular structure will be solved after the molecular dynamics simulation system runs for a short period of time. At a certain temperature and pressure, the coordinates of the molecular dynamics simulation system will be written to the output file. By calculating in this way, a series of coordinates that change with time can be obtained, which is the motion trajectory of the molecular structure at different temperatures and different times. By analyzing the obtained trajectory, many macroscopic properties of the simulation system can be extracted from the output file.

[0063] The velocity Verlet algorithm in GROMACS is used to solve the equation of motion of the above molecular structure. In the velocity Verlet algorithm, the position r and velocity v of the molecular structure at time t are used to integrate and solve the equation of motion.

[0064]

[0065] Given a starting file with the initial position x(0) and initial velocity v(0) of the molecular structure, the Verlet algorithm is used to solve equations (3) and (4). Verlet integration is a calculation method that integrates the past, present, and future, with an accuracy of O(4), good stability, and low algorithm complexity.

[0066] Some basic settings of the molecular dynamics simulation system include: simulation at constant temperature and constant pressure, and the cutoff distance of electrostatic interaction is The cutoff distance for the Lennard-Jones interaction is also The bond lengths of the solutes were constrained using the LINCS algorithm. A neighboring grid search method was used and updated every ten steps. The equations of motion were calculated using the Verlet algorithm. The time step was 2f s , coordinates every 2p sSave once. The run time is 10μs, and the simulation temperatures are 290K, 360K, 370K, 380K, 390K and 400K. At 290K, the system is minimized for 20ns, and then the initial structure of the simulation is selected from the conformation. Due to the different lengths of the two loops in the L1 region and the L2 region, the melting temperatures of the bases in the Stem region are also different.

[0067] Finally, the field force is set. The field force is mainly composed of the potential energy U (r1, r2, ..., r N ) and a series of parameters. These parameters are usually obtained from abinitio or semi-empirical quantum mechanical calculations or by fitting experimental neutron, X-ray and electron diffraction, nuclear magnetic resonance, infrared, Raman and neutron spectroscopy.

[0068] By setting the periodic boundary conditions, motion equations, field forces, temperature, pressure, and running time of the molecular dynamics simulation system, and running the molecular dynamics simulation system, the motion trajectory of the molecular structure at different temperatures and times can be obtained.

[0069] It should be noted that periodic boundary conditions (PBC) are a set of boundary conditions, which are usually used in small units to simulate approximate large (infinite) systems. PBC is usually used for computer simulation and mathematical models. For finite systems, setting periodic boundary conditions can reduce edge effects. The atoms of the system to be simulated are placed in a space-filling box, and the atoms in the present embodiment are several specific bases or base pairs in the loop region / Stem region in the RNA pseudoknot structure. The periodic boundary conditions simply copy the fields on one side, such as the left side of the region, to the right side of the region. This type of boundary condition makes it possible to simulate the entire molecular system by simulating only a small part of the atoms. When the physical geometry of the object of study and the expected flow pattern and thermal effect have periodic repetitive properties, periodic boundary conditions can be used.

[0070] S2: Based on the motion trajectory, the geometric parameters of specific bases in the loop region / Stem region of the molecular structure during the open-closed state transition are obtained. The geometric parameters include the RMSD of the base, the torsion angle, and the dihedral angle formed by each specific base.

[0071] In one embodiment, in order to obtain the thermodynamic parameters of the pseudoknot more accurately, the present invention studies a typical pseudoknot structure. The simplest RNA pseudoknot is the H-type pseudoknot (e.g. Figure 3), a simple pseudoknot is composed of two helices forming a stem and two loops spanning the helical stem. RNA pseudostructure is defined as the pairing between a structural loop with bases and other regions of RNA. In addition, the secondary structure information of RNA is a key step in predicting its tertiary structure. The essence of predicting RNA secondary structure is to find out the pairing of bases in the Stem region in the secondary structure and the thermodynamic properties of the loop region on the basis of the primary structure. For this purpose, the present application uses all-atom dynamics simulation to study the influence of the loop region in the H-type pseudoknot on the thermodynamic and kinetic properties of the Stem region.

[0072] In this application, the S2 region and L1 region in the H-type pseudoknot structure are first selected to study the influence of the loop region on the thermodynamic parameters and kinetic characteristics of the Stem region.

[0073] by Figure 2 Taking the structure of HIV (PDB ID: 1hvu) as an example, the specific bases selected are residues 8, residues 27, residues 6 and residues 7. Except for the atoms on residues 8, residues 27, residues 6 and residues 7, the atoms of other residues on the nucleotide are fixed by using additional forces (force constant: 1000 kJ / mol nm -2 ). In order to accelerate the transition between the closed state and the open state, high temperature simulation is used to simulate the entire system. The selected simulation temperatures are 360K, 370K, 380K, 390K and 400K, and each simulation temperature lasts for 10μs, that is, the total simulation time at each temperature is 10μs.

[0074] The running trajectory of the specific base at each temperature is obtained. This trajectory can present the transition process between the open and closed states of the base pair, wherein the position of the specific base under different states is obtained by tracking the speed and position of the specific base. In this embodiment, the RMSD (root mean square deviation) of the two specific bases U8 and the specific base 27 in the S2 region, and the dihedral angle formed by the specific base U8 and the specific base U9 are calculated respectively. Among them, the dihedral angle formed by the four atoms in the specific base U8 and the specific base U9 is C1'(i)-N1(i)-C1'(i+1)-N1(i+1). The relationship between the RMSD of the specific base U8 and the simulation time is as follows: Figure 4 As shown, the dihedral angle formed by the specific base U8 and the specific base U9 is as follows Figure 5 According to the simulation trajectory, the RMSD and torsion angle of the specific base 27 were calculated. It should be noted that the torsion angle can be divided into non-ring torsion angle, equilibrium torsion angle and phosphate backbone torsion angle. The torsion angle here is the phosphate backbone torsion angle C3'(i)-O3'(i)-P(i+1)-O5'(i+1).

[0075] Optionally, obtaining geometric parameters of specific bases in the loop region / Stem region in the molecular structure during the open-closed state transition based on the motion trajectory includes: counting the positions of each specific base in the loop region / Stem region in the molecular structure at different times; and calculating the RMSD, torsion angle and dihedral angle of each specific base according to the position of each specific base.

[0076] In one embodiment, based on solving the trajectory of the specific base, the RMSD, torsion angle and dihedral angle formed by each specific base at any time during the movement are determined according to the position of each specific base. For example, the RMSD of the specific base U8 is calculated, such as Figure 4 As shown, when the RMSD is about When RMSD is greater than When the specific base U8 flips into the solvent, and sometimes the specific base U8 falls into the large groove, and this state lasts for a long time. The dihedral angle formed by the specific base U8 and the specific base U9 can also be calculated through the simulation trajectory, C1'(i)-N1(i)-C1'(i+1)-N1(i+1), such as Figure 5 As shown. When the central region of the dihedral angle is -160°, the specific base U8 remains in a folded state and vibrates near the initial position. In this state, the RMSD corresponding to the conformation of the specific base U8 is ~ When the dihedral angle is at 60° in the center, the specific base U8 is thrown into the solvent and sometimes stuck in the groove for a long time. In this state, the RMSD of the corresponding conformation is greater than

[0077] S3: Determine the RMSD, torsion angle, and dihedral angle ranges of specific bases at different times to determine the state of the specific bases.

[0078] Optionally, determining the range of two-state conditions satisfied by the RMSD, torsion angle, and dihedral angles of specific bases at different times to determine the state of the specific bases includes: if the RMSD region of a specific base constituting a base pair at any time is between The RMSD region of another specific base is ~ When the dihedral angle formed by the two specific bases ranges from ∼-80° to ∼-140°, and the torsion angle is between ∼-50° and ∼-100°, the state of the two specific bases is determined to be a closed state.

[0079] In one embodiment, the closed state condition range is: the RMSD region of a specific base of the base pair is ~ The RMSD region of another specific base is ~ The dihedral angle formed by two specific bases ranges from ∼-80° to ∼-140°, and the torsion angle is between ∼-50° and ∼-100°. Specifically, at a random moment, the RMSD of the specific base U8 is ∼ And the dihedral angle is ~-80° to -140°. It is also necessary to meet the specific base 27, RMSD is ~ And the torsion angle is between -50° and -100°. In this state, the specific base U8 and the specific base 27 only vibrate at their initial positions, and the base pairing and base stacking between them are complete and not destroyed. At this time, it can be judged that the specific base pairing and base stacking of the S2 region are complete and in a closed state, always maintaining 6bp, such as Figure 7-8 As shown, Figure 6 For the simulation temperature at 360K and the total simulation time of 10μs, the RMSD of the specific base 27 changes with the simulation time; Figure 7 The distribution of RMSD of specific base 27 during the whole simulation process for the simulated temperature of 360K; Figure 8 Distribution of phosphate backbone torsion angles for a specific base 27 during the entire simulation for a simulated temperature of 360 K.

[0080] Optionally, determining the range of two-state conditions satisfied by the RMSD, torsion angle, and dihedral angles of the specific bases at different times, and determining the state of the specific bases further includes:

[0081] If the RMSD of a specific base constituting a base pair at any time is greater than Another specific base has an RMSD greater than When the dihedral angle formed by the two specific bases is between -140° and 100°, and the torsion angle is between 50° and 100°, the state of the two specific bases is determined to be the open state.

[0082] In one embodiment, the condition range of the open state is: the RMSD of a specific base constituting the base pair is greater than The dihedral angle formed by two specific bases is between -140° and 100°; or the RMSD of another specific base is greater than And the torsion angle is between 50° and 100°. At a random moment, the RMSD of a specific base U8 is greater than And the dihedral angle is between -140° and 100°, the specific base is 27, and the RMSD is greater than And the torsion angle is between 50° and 100°. If one or both of these two conditions are met, the conformation is defined as the open state. In this state, the base pair and base stacking are destroyed when one of the specific bases U8 and the specific base 27 is thrown into the solvent. And at least one base is thrown into the solvent.

[0083] S4: The average lifetime, number of times, and probability of a specific base in the closed and open states during the total time of the state statistics simulation of the specific base.

[0084] Optionally, the average lifetime, number and probability of a specific base in a closed state and an open state during the total time of a statistical simulation of the state of the specific base include: separately counting the number of times each specific base appears in a closed state or an open state at each simulation temperature within the total time of each simulation; calculating the average lifetime and probability of the closed state according to the number of closed states and the time each time the closed state is maintained, or calculating the average lifetime and probability of the open state according to the number of open states and the time each time the open state is maintained.

[0085] In one embodiment, as shown in Table 1, Table 1 shows the average lifespan, number of times and probability of the closed state and the open state when the total simulation time is 10 μs and the simulation temperatures are 360K, 370K, 380K, 390K and 400K respectively.

[0086] Table 1

[0087]

[0088] in, is the average lifetime in the closed state, is the average lifetime in the open state, N c N is the number of times the conformation appears in the closed state during the total simulation time. o is the total number of times the conformation appears in the open state during the total simulation time, p o is the probability of the open state at simulated temperature T, p c is the probability of the closed state at the simulated temperature T.

[0089] At each simulation temperature, τ is the total simulation time, τ = 10 μs, and the probability of maintaining the closed state during the total simulation time is: in, is the time period of the ith stay in the closed state. The probability of remaining in the open state during the total simulation time is: in, is the time period of the i-th stay in the open state.

[0090] like Fig. 9As shown in the figure, the probability of the closed state during the simulation at all simulation temperatures is shown. It can be seen that the probability of the closed state for all simulation temperatures with a simulation time of 9600ns reaches a stable value, indicating that the transition balance between the open and closed states has been achieved. In order to ensure that the simulation reaches the closed-open transition balance, each simulation temperature lasts until 10μs. Fig.10 As shown, It is linearly related to the inverse of temperature (1 / T).

[0091] The average lifetime of the open and closed states can be obtained as follows: where N is the total number of times a particular base is present in one state. The average lifetimes of the open and closed states at different simulation temperatures are given by Fig.11 As shown, Fig.12 The lifetime distribution of the open and closed states is shown. The average lifetime of the open state and the average lifetime of the closed state show different correlation behaviors with temperature. The simulation temperature has little effect on the average lifetime of the open state, and it is weakly correlated with the simulation temperature. The average lifetime of the closed state is more affected by temperature, and it is strongly correlated with the simulation temperature.

[0092] S5: The thermodynamic parameters and kinetic parameters of the specific base are calculated based on the average lifespan, number and probability of the specific base in the closed state and the open state, wherein the thermodynamic parameters include the free energy difference and the linear relationship between the free energy difference and the temperature; the kinetic parameters include the closing rate and the opening rate.

[0093] Optionally, calculating the thermodynamic parameters and kinetic parameters of a specific base based on the average lifetime, number and probability of a specific base in a closed state and an open state includes: calculating the thermodynamic parameter free energy difference of base pairs formed by the specific base at different temperatures based on the enthalpy variable and entropy variable of the transition between the closed state and the open state of the specific base.

[0094] The linear relationship between the free energy difference and temperature was calculated based on the probability of the closed and open states of a specific base.

[0095] Based on the average lifetime and number of closed / open states, the kinetic parameters closing rate / opening rate of the specific base are calculated respectively.

[0096] In one embodiment, according to the statistical data in Table 1, since the simulated conformations are divided into open and closed states, the thermodynamic parameters of the specific bases are first calculated, and the probabilities of the closed and open states are used to participate in the calculation of the following formula (5) to obtain the closing free energy difference ΔG between the specific base 8 and the specific base 27.

[0097]

[0098] Among them, k Bis the Boltzmann constant, It represents the linear relationship between the free energy difference ΔG and the temperature T.

[0099] ΔG = ΔH-TΔS, where ΔH is the enthalpy change of the transition between the closed state and the open state, and ΔS is the entropy change of the transition between the closed state and the open state. The thermodynamic parameters of the AU base pairs in the Stem region of the RNA pseudoknot were calculated to be ΔH = -7.69 kcal / mol and ΔS = -21.46 eu. In addition, from Fig.10 As can be seen from the figure, the melting temperature is 358K, while for the same terminal base pair without pseudoknot, they are ΔH = -7.3lcal / mol and ΔS = -18.5eu, and the melting temperature is 395K. Therefore, the existence of the Loop region reduces the melting temperature of the Stem region. And the enthalpy change and entropy change become larger.

[0100] Secondly, the kinetic parameters of the specific bases were calculated. For the two-state transition from open to closed, the closing rate k of the specific base 8 and the specific base 27 in the Stem region was + and the opening rate k - The average lifetime of the open and closed states can be obtained as Compared with the average lifetime of the open and closed states of a single base pair, it is found that at the same temperature, the average lifetime of the closed state becomes smaller, while the average lifetime of the open state becomes larger. This shows that the presence of the loop region does not change the opening rate k - , and the closing rate k + Become smaller.

[0101] The present application also provides a device for obtaining the thermodynamic and kinetic parameters of RNA pseudoknot structures, such as Fig.13 As shown, it includes: a motion trajectory acquisition module, which is used to determine the molecular structure, and simulate the motion trajectory of the molecular structure at different temperatures and different times based on molecular dynamics simulation software. The motion trajectory includes the speed and position of the molecular structure at different times.

[0102] The geometric parameter calculation module is used to obtain the geometric parameters of specific bases in the loop region / Stem region in the molecular structure during the open-closed state transition based on the motion trajectory. The geometric parameters include the RMSD of the base, the torsion angle, and the dihedral angle formed by each specific base.

[0103] The state determination module is used to determine the RMSD, torsion angle and dihedral angle range of specific bases at different times and determine the state of the specific bases.

[0104] The calculation module is used to calculate the average lifespan, number of times and probability of a specific base in a closed state and an open state during the total time of a statistical simulation of the state of the specific base.

[0105] The output module is used to calculate the thermodynamic parameters and kinetic parameters of the specific base according to the average lifespan, number and probability of the specific base in the closed state and the open state, wherein the thermodynamic parameters include the free energy difference and the linear relationship between the free energy difference and the temperature; the kinetic parameters include the closing rate and the opening rate.

[0106] It should be noted that the method that can be implemented by the device for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure provided in the present application is completely consistent with the method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure described above, and will not be repeated here.

[0107] In one embodiment, in another embodiment of the present application, a terminal device is provided, the terminal device includes a processor and a memory, the memory is used to store a computer program, the computer program includes program instructions, and the processor is used to execute the program instructions stored in the computer storage medium. The processor may be a central processing unit (CPU), or other general-purpose processors, digital signal processors (DSP), application-specific integrated circuits (ASIC), field-programmable gate arrays (FPGA) or other programmable logic devices, discrete gates or transistor logic devices, discrete hardware components, etc., which are the computing core and control core of the terminal, which are suitable for implementing one or more instructions, and are specifically suitable for loading and executing one or more instructions to implement the corresponding method flow or corresponding functions; the processor described in the embodiment of the present application can be used for the operation of the method for obtaining the thermodynamic and kinetic parameters of the RNA pseudoknot structure.

[0108] In one embodiment, in another embodiment of the present application, the present application also provides a storage medium, specifically a computer-readable storage medium (Memory), which is a memory device in a terminal device for storing programs and data. It is understandable that the computer-readable storage medium here can include both the built-in storage medium in the terminal device and the extended storage medium supported by the terminal device. The computer-readable storage medium provides a storage space, which stores the operating system of the terminal. In addition, one or more instructions suitable for being loaded and executed by the processor are also stored in the storage space, and these instructions can be one or more computer programs (including program codes). It should be noted that the computer-readable storage medium here can be a high-speed RAM memory or a non-volatile memory, such as at least one disk memory. The processor can load and execute one or more instructions stored in the computer-readable storage medium to implement the method for obtaining the thermodynamic and kinetic parameters of the RNA pseudoknot structure in the above embodiment.

[0109] Those skilled in the art will appreciate that the embodiments of the present application may be provided as methods, systems, or computer program products. Therefore, the present application may adopt the form of a complete hardware embodiment, a complete software embodiment, or an embodiment in combination with software and hardware. Moreover, the present application may adopt the form of a computer program product implemented in one or more computer-usable storage media (including but not limited to disk storage, CD-ROM, optical storage, etc.) that include computer-usable program code.

[0110] The present application is described with reference to flowcharts and / or block diagrams of methods, devices (systems) and computer program products according to embodiments of the present application. It should be understood that each process and / or box in the flowchart and / or block diagram, as well as the combination of processes and / or boxes in the flowchart and / or block diagram, can be implemented by computer program instructions. These computer program instructions can be provided to a processor of a general-purpose computer, a special-purpose computer, an embedded processor or other programmable data processing device to generate a machine, so that the instructions executed by the processor of the computer or other programmable data processing device generate instructions for implementing the processes in the flowchart and / or block diagram. Figure 1 A process or multiple processes and / or boxes Figure 1 A device that provides the functions specified in a block or multiple blocks.

[0111] These computer program instructions may also be stored in a computer readable memory capable of directing a computer or other programmable data processing device to operate in a specific manner, so that the instructions stored in the computer readable memory produce an article of manufacture including an instruction device, which implements the process Figure 1 A process or multiple processes and / or boxes Figure 1 A function specified in one or more boxes.

[0112] These computer program instructions can also be loaded onto a computer or other programmable data processing device so that a series of operating steps are executed on the computer or other programmable device to produce a computer-implemented process, thereby providing instructions for implementing the process in the computer or other programmable device. Figure 1 A process or multiple processes and / or boxes Figure 1 The steps for the functions specified in one or more boxes.

[0113] Finally, it should be noted that the above embodiments are only used to illustrate the technical solution of the present application rather than to limit it. Although the present application has been described in detail with reference to the above embodiments, ordinary technicians in the relevant field should understand that the specific implementation methods of the present application can still be modified or replaced by equivalents, and any modifications or equivalent replacements that do not depart from the spirit and scope of the present application should be included in the scope of protection of the claims of the present application.

Claims

1. A method for obtaining thermodynamic and kinetic parameters of RNA pseudoknot structure, characterized in that: include: Determine the molecular structure, and simulate the motion trajectory of the molecular structure at different temperatures and at different times based on molecular dynamics simulation software; Based on the motion trajectory, the geometric parameters of the specific bases in the loop region / Stem region in the molecular structure during the open-closed state conversion process are obtained, wherein the geometric parameters include the RMSD, the torsion angle, and the dihedral angle formed by each specific base of the base; Determine the state of the specific base by determining the RMSD, torsion angle, and dihedral angle of the specific base at different times and the range of the two-state conditions satisfied by the dihedral angles formed by each specific base; The average lifespan, number of times and probability of the specific base in the closed state and the open state during the total time of the state statistical simulation of the specific base; The thermodynamic parameters and kinetic parameters of the specific base are calculated according to the average lifespan, number of times and probability of the specific base in the closed state and the open state.

2. The method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure according to claim 1, characterized in that: Determine the molecular structure, and simulate the motion trajectory of the molecular structure at different temperatures and times based on molecular dynamics simulation software, including: Download the molecular structure from the PDB library and import it into a molecular dynamics simulation system; The periodic boundary conditions, motion equations, field forces, temperature, pressure, and running time of the molecular dynamics simulation system are set, and the molecular dynamics simulation system is run to obtain the motion trajectories of the molecular structure at different temperatures and at different times.

3. The method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure according to claim 2, characterized in that: The molecular dynamics simulation system is run to obtain the motion trajectories of the molecular structure at different temperatures and at different times, including: Select the molecular dynamics simulation mode, set the temperature, pressure, and total simulation time; Given the initial position and initial velocity of a specific base, the velocity Verlet algorithm is used to solve the Newton's motion equations in the molecular dynamics simulation mode to obtain the motion trajectory of the molecular structure at different temperatures and times.

4. The method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure according to claim 1, characterized in that: The geometric parameters of a specific base in the loop region / Stem region of the molecular structure during the open-closed state conversion process are obtained based on the motion trajectory, including: Count the positions of specific bases in the loop region / Stem region of the molecular structure at different times; According to the position of each of the specific bases, the RMSD, torsion angle, and dihedral angle formed by each specific base are calculated.

5. The method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure according to claim 1, characterized in that: Determining the range of two-state conditions satisfied by the RMSD, torsion angle, and dihedral angles of the specific bases at different times, and determining the state of the specific bases includes: If the RMSD region of a specific base constituting a base pair at any time is Another specific base RMSD region is When the dihedral angle formed by the two specific bases ranges from -80° to -140°, and the torsion angle is between -50° and -100°, the state of the two specific bases is determined to be a closed state.

6. The method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure according to claim 1, characterized in that: Determining the range of two-state conditions satisfied by the RMSD, torsion angle, and dihedral angles of the specific bases at different times, and determining the state of the specific bases further includes: If the RMSD of a specific base constituting a base pair at any time is greater than Another specific base has an RMSD greater than When the dihedral angle formed by the two specific bases is between -140° and 100°, and the torsion angle is between 50° and 100°, the state of the two specific bases is determined to be an open state.

7. The method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure according to claim 1, characterized in that: The average lifespan, number of times and probability of the specific base in the closed state and the open state during the total time of the state statistical simulation of the specific base include: The number of times each specific base appears in the closed state or the open state in the total time of each simulation at each simulation temperature is counted separately; The average life span and probability of the closed state are calculated according to the number of closed states and the time each time the closed state is maintained, or the average life span and probability of the open state are calculated according to the number of open states and the time each time the open state is maintained.

8. The method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure according to claim 1, characterized in that: The thermodynamic parameters and kinetic parameters of the specific base are calculated based on the average lifespan, number of times and probability of the specific base in the closed state and the open state, including: Calculating the free energy difference of thermodynamic parameters of base pairs formed by the specific bases at different temperatures according to the enthalpy variable and entropy variable of the transition between the closed state and the open state of the specific base; Calculating a linear relationship between the free energy difference and the temperature based on the probability of the closed state and the open state of the specific base; Based on the average lifetime and number of closed / open states, the kinetic parameters closing rate / opening rate of the specific base are calculated respectively.

9. A device for obtaining thermodynamic and kinetic parameters of RNA pseudoknot structure, characterized in that: include: A motion trajectory acquisition module is used to determine the molecular structure and simulate the motion trajectory of the molecular structure at different temperatures and at different times based on molecular dynamics simulation software. The motion trajectory includes the speed and position of the molecular structure at different times. A geometric parameter calculation module, for obtaining geometric parameters of specific bases in the loop region / Stem region of the molecular structure during the open-closed state conversion process based on the motion trajectory, wherein the geometric parameters include RMSD, torsion angle, and dihedral angle formed by each specific base of the base; A state determination module, used to determine the RMSD, torsion angle and dihedral angle ranges of the specific bases at different times, and determine the state of the specific bases; A calculation module, for calculating the average lifespan, number of times and probability of the specific base in the closed state and the open state during the total time of the state statistical simulation of the specific base; An output module is used to calculate the thermodynamic parameters and kinetic parameters of the specific base according to the average lifespan, number and probability of the specific base in the closed state and the open state, wherein the thermodynamic parameters include the free energy difference and the linear relationship between the free energy difference and the temperature; the kinetic parameters include the closing rate and the opening rate.

10. A computer-readable storage medium storing a computer program, characterized in that: When the computer program is executed by a processor, the method for obtaining thermodynamic and kinetic parameters of an RNA pseudoknot structure as claimed in any one of claims 1 to 8 is implemented.

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