Peptides causing formation of compact structure

A member, said technology, applied in the field of peptides causing the formation of compact structures, able to solve problems such as difficulties

Inactive Publication Date: 2001-07-04
RIGEL PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although, as is known in the art, linear peptides with freely rotating amino and carboxy termini can be powerful, converting such peptide structures into pharmacological agents is often difficult

Method used

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  • Peptides causing formation of compact structure
  • Peptides causing formation of compact structure
  • Peptides causing formation of compact structure

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0169] Novel peptides that form observable dimers under harsh conditions

[0170] By perfusing 3 x 10 -4 M, the EFLIVKS-amide solution of pH 6.4 is entered in the Finnigan LCQ ion trapping mass spectrometer of electrospray source, and this peptide appears to self-associate to form dimer ( Figure 3A ), after a temperature of 210 °C in the inlet capillary and harsh electrospray conditions, the molecular weight was detected in the gas phase to be exactly twice the molecular weight of the monomer, so it is expected to dimerize at significantly lower concentrations in aqueous solution. This peptide also forms a dimer (also detectable by mass spectrometry) when eluted from a C18 reverse phase column at about 25% acetonitrile at about pH 2.5 ( Figure 3B ). When both are continuously infused into the ion trap mass spectrometer through the electrospray interface, the comparison Figure 3A and detect the peptide SKVILFE (it is in 10 -13 M in aqueous solution to form dimers (Bodenm...

Embodiment 2

[0173] Example 2 When adding 18mer detection peptide N- and C-terminal

[0174] EFLIVKS forms compact proteolysis-resistant structures

[0175] When fused to the N- and C-termini of the 18mer detection polypeptide, the peptide EFLIVKS can form a compact structure of this polypeptide (also referred to herein as peptide 1). The 18mer polypeptide sequence is VGTIVTMEYRIDRRTRSFV from barley c2-chymotrypsin inhibitor [Leatherbarrow and Salacinski, Biochemistry 30:10717-21 (1991)]. This peptide analog, with cysteines substituted for valines at both the N- and C-termini, is thought to fold into a compact structure similar to the loop in chymotrypsin-2. Such a compact structure should be a poor substrate for proteases such as elastase, and has indeed been suggested as an inhibitor of both variants of elastase, chymotrypsin and subtilisin. This disulfide cyclized analog has been synthesized and tested by us and is actually a poor protease substrate, but still ...

Embodiment 3

[0177] Detection of low-energy conformers of peptide 1

[0178] To examine the structural properties of the compact structure of peptide 1, low energy conformers were obtained using high temperature molecular dynamics simulated annealing methods similar to those published in Nilges et al., Protein Engineering 2:27-38 (1988), as performed in Discover95. Structure saved every 2psec (from 900K, different trajectories with duration 400-1400psec), cooled to 300K with 5ps time, and distance-dependent dielectric constant (ε=80 at 80 angstrom interval to ε=1 at 1 angstrom interval linear variation between ) with 200 steps of the steepest descent algorithm for minimization, and then use as many steps as necessary with the gradient algorithm to give the largest amount of derivatives less than 0.001 kcal / A. Collect and compare the resulting low-energy structures from orbitals starting from the following structures:

[0179] a) Peptide 1, the 18mer polypeptide derived fro...

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Abstract

The present invention relates to compositions and methods comprising peptides with high mutual affinity that, when attached to a protein, assist the protein to fold into a compact structure. By virtue of its stability and binding, this scaffold extends the activity of any contained protein sequence in the presence of cellular and other proteases. This compact structure may have other included functional sequences, which are superior to linear and less constrained peptides for library screening, building structure-biased peptide libraries, and targeting specific intracellular and extracellular compartments. The compositions of the present invention can be displayed on viral, archaeal, prokaryotic and eukaryotic cell surfaces for library screening, drug screening and display. The methods of the present invention are useful for in vivo screening of intracellular effector proteins that modulate signaling pathways, and for identification of interacting proteins in vitro. Therefore, the present invention can be used as a scaffold for gene therapy, for the isolation of new therapeutic drugs, and has potential utilization value for use as a therapeutic agent in physiological fluids.

Description

field of invention [0001] The compositions and methods of the invention involve the use of dimerization peptides that can self-associate and their use with other proteins to induce compact structure formation. Background of the invention [0002] Proteins mostly interact through conformationally restricted domains. Although, as is known in the art, linear peptides with freely rotating amino and carboxy termini can be powerful, converting such peptide structures into pharmacological agents is often difficult. Displaying peptides in conformationally constrained structures can thus yield drugs with high affinity for their target proteins. Constrained peptides have a number of useful features compared to their linear analogs. They include: (i) increased stability to proteolytic hydrolysis due to the absence of unconstrained N- or C-terminal amino acids susceptible to aminopeptidases or carboxypeptidases and a non-extended structure that reduces susceptibility to endopeptidases...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/09C07K1/04C07K7/06C07K14/00C07K14/47C12N1/15C12N1/19C12N1/21C12N5/10C12P21/02C12Q1/02C12Q1/68
CPCC07K7/06C07K14/47C07K1/047
Inventor D·安德森T·格鲁拉加
Owner RIGEL PHARMA
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