10 results about "Thermostable enzymes" patented technology

The application discloses a kind of high thermal stability glutamine transaminase mutant and its method of expression in streptomyces, belong to enzyme engineering field.The high thermal stability glutamine transaminase mutant provided by the application can efficiently catalyze casein crosslinking at high temperature, realizes the heterologous efficient expression of glutamine transaminase.Rational mutation is carried out to key amino acid site of glutamine transaminase, and the mutant strain with significantly improved thermal stability is obtained.Among them, the optimal mutant strain FRAPD-TGm2A3 has a half-life of up to 537.91 min at 60 DEG C.The mutant can still efficiently catalyze the crosslinking reaction of casein at high temperature of 85 DEG C.The present application not only lays a solid research foundation for the modification of thermal stability of glutamine transaminase, but also provides a generalizable strategy framework for the rational modification of other thermostable enzymes, and has important industrial application value and academic reference significance.

Provided herein are secretory compositions / cocktails derived from a novel, extremophilic fungal species cultured, isolated, and characterized herein. Said novel, extremophilic fungal species may be exploited to produce secretory compositions / cocktails which comprise a unique mixture of thermo-tolerant, stable enzymes capable of digesting cellular and genetic material in its surroundings under extreme temperatures. Such secretory compositions / cocktails may be useful as cleaning and sterilization solutions against biofilm, urine, yeast, bacterial, or viral contamination. Other uses include, but are not limited to, urea removal, fertilizer wastewaters, wastewater reclamation for life support systems in space, pH control, source of ammonia and carbon dioxide, starch digestion, biofuel production, as pharmaceuticals, and medical treatments such as wounds, skin disorders, or nail disorders.

The present invention discloses a method for designing and modifying a new thermostable enzyme based on a deep learning model. By fusing a protein thermostability discrimination model and a protein sequence generation model, a new enzyme sequence with thermostability characteristics is directly generated. The present invention utilizes the optimal growth temperature information of heat-resistant species and applies a deep learning model to extract the thermostable characteristics of heat-resistant species, avoiding the defects of insufficient heat-resistant protein data and the bias caused by manual feature extraction; utilizing the protein sequence generation model, the protein sequence space can be effectively expanded; the two model data feedback loops directly generate new enzyme sequences with thermostable characteristics, which can overcome the path dependence problem of traditional directed evolution methods and significantly reduce the size of the directed evolution screening library; utilizing the present invention, new enzyme sequences with thermostable characteristics can be efficiently obtained.

This invention discloses a strategy for rapid detection of isoniazid resistance in Mycobacterium tuberculosis based on multiplex polymerase chain reaction (Multiplex PCR) using local rTth DNA polymerase and interpretation of amplification patterns. This strategy utilizes a specific primer combination that allows simultaneous amplification of the control fragment and the target fragment of the katG gene codon 315 mutation in a single PCR reaction. The amplification products are analyzed by 1% (w / v) agarose gel electrophoresis and interpreted based on the resulting DNA banding pattern to distinguish sensitive isolates, isolates carrying the katG315 mutation causing isoniazid resistance, and invalid test results. The use of local rTth DNA polymerase provides an alternative thermostable enzyme to support national diagnostic raw material independence without changing the detection principle.This strategy allows for rapid identification of katG315 mutations associated with isoniazid resistance without the need for DNA sequencing or further molecular analysis. The invention provides a simple, rapid, specific, easily interpretable, and cost-effective molecular diagnostic method to support the early detection of isoniazid-resistant tuberculosis.

The invention provides a Pfu-MutS collaborative error correction PCR method and application thereof, and relates to the technical field of gene engineering. According to the method, high-fidelity Pfu DNA polymerase and heat-stable TaqMutS enzyme are synchronously introduced into a PCR reaction system, a mismatch-containing DNA chain is combined in real time through the mismatch recognition capability of MutS and the extension of the mismatch-containing DNA chain is inhibited, and meanwhile, mismatch bases are corrected by utilizing the activity of 3 '-> 5' exonuclease of Pfu, so that a dual error correction mechanism is formed. Complex protein fusion design is not needed, real-time error correction is achieved by directly utilizing the combination of commercially available independent enzymes, the development cost and the operation complexity are remarkably reduced, meanwhile, the additional purification step needed by traditional step-by-step error correction is omitted, and the method is suitable for high-fidelity requirements of conventional PCR amplification, long-fragment synthesis and high-GC-content templates and has wide application prospects. And an efficient and economic general solution is provided for the fields of DNA synthesis and gene editing.

The disclosure provides a thermostable enzyme-linked biosensor expression cassette comprising a nucleic acid comprising a nucleotide sequence encoding a β-glucosidase reporter. The enzyme-linked biosensor expression cassette of the disclosure comprises a nucleic acid comprising a nucleotide sequence encoding (i) a transcription factor, (ii) a promoter, (iii) a terminator, and (iv) a cloning site for a gene of interest. The disclosure further provides novel variants of β-glucosidase that function as the reporter enzyme and exhibit superior properties (e.g., without limitation, pH stability and thermal stability) compared to existing β-glucosidase, providing improved biosensor expression cassettes.

Novel enzyme variants of naturally occurring wild-type Thermosyntropha lipolytica Tl_Est47 are provided, which have improved thermotolerance properties that do not adversely affect the stability or activity of the enzyme variants over a range of temperatures or pH values.

The invention discloses an L-threonine aldolase mutant with high thermal stability and a method for synthesizing droxidopa by using the L-threonine aldolase mutant, and belongs to the field of biological catalysis engineering. The L-threonine aldolase mutant with high thermal stability provided by the invention can be used for efficiently catalyzing the generation of droxidopa, so that the heterologous efficient expression of the L-threonine aldolase is realized. Reasonable mutation is carried out on key amino acid sites of the L-threonine aldolase through strategies such as sequence conservative analysis, solvation free energy prediction and folding free energy prediction, and a mutant strain with remarkably improved thermal stability is obtained. Wherein the yield of droxidopa catalyzed by the mutant strain L17F / V12R / L206W with the optimal thermal stability in a 1L system at the temperature of 35 DEG C is 5.52 g / L. The invention not only lays a solid research foundation for high-efficiency industrial preparation of droxidopa by a biological method, but also provides a generalizable strategy framework for rational transformation of thermostable enzyme, and has important industrial application value and academic reference significance.

The invention provides a reversibly modified thermostable enzyme as well as a preparation method and application thereof, and relates to the technical field of biology. The reversibly modified thermostable enzyme is formed by non-covalent binding of at least two thermostable enzyme modifiers and a thermostable enzyme, the activity of the modified enzyme is inhibited in a wider temperature range, and the defect of the thermostable enzyme modified by a single thermostable enzyme modifier is relieved.

The invention relates to the technical field of thermostable enzymes, in particular to beta-1, 3-glucosidase and a method for improving the enzyme activity of the beta-1, 3-glucosidase. The beta-glucosidase encoding gene YALI1B03564g is identified in a yarrowia lipolytica genome for the first time and is named as beta-1, 3-glucosidase, and the beta-glucosidase encoding gene YALI1B03564g is confirmed to have unique beta-1, 3-glucosidic bond substrate specificity through heterologous expression and enzymatic characterization; the thermal stability of the enzyme is further remarkably improved through rational design, and the enzyme is applied to in-vitro polysaccharide degradation; the technical scheme provided by the invention further lays a foundation for large-scale production of the functional yeast beta-glucan oligosaccharide.