Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Glutamine transaminase with improved enzymatic activity and thermal stability

A technology of glutamine and thermostability, which is applied in the field of transglutaminase, can solve problems such as poor thermostability, low specific enzyme activity, and lack of enzymes, and achieve the effect of reducing production costs and improving production efficiency

Inactive Publication Date: 2013-12-11
JIANGNAN UNIV
View PDF1 Cites 4 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In order to improve the current situation of low specific enzyme activity and poor thermal stability of MTG, this study deleted the N-terminal amino acid of MTG, thereby reducing the binding resistance between the substrate and the active center of MTG, and improving the affinity of MTG for the substrate.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Glutamine transaminase with improved enzymatic activity and thermal stability
  • Glutamine transaminase with improved enzymatic activity and thermal stability
  • Glutamine transaminase with improved enzymatic activity and thermal stability

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1: Simulation of MTG crystal structure derived from Streptomyces hygroscopicus

[0020] Using the reported TGase crystal structure of S.mobaraensis as a template, simulate the crystal structure of S.hygroscopicus TGase on the swiss-model website (http:∥swissmodel.expasy.org / ), as shown in figure 1 shown.

Embodiment 2

[0021] Example 2: Obtainment of highly active mutant strains (Del1-4)

[0022] 1. Use the site-directed mutagenesis kit (TaKaRa) to delete 7 amino acids (Asp 1, Ala 2, Ala 3, Asp 4, Glu 5, Arg 6, Val 7) at the N-terminus, and delete one amino acid named Del 1. By analogy, the deletion of the first seven amino acids was named Del1-7, and the transformant was sequenced by Shanghai Sangong.

[0023] 2. Transform the plasmid with correct sequencing into E.coli BL 21, select the transformant and inoculate it into LB liquid medium, culture at 37°C for 12 hours, and transfer it to TB medium with an inoculation volume of 3%. Bacteria grow to OD 600 When it was 2, IPTG was added to induce, and the culture temperature was lowered to 20°C, and cultured for 48h.

[0024] 3. Collect the fermentation supernatant, detect the enzyme activity of the fermentation supernatant, and purify the sample on a His-nickel column. The purified protein electrophoresis is as follows: figure 2 shown.

...

Embodiment 3

[0028] Example 3: Obtainment of highly active and thermostable mutants (TG-Del 1-4E5D)

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention provides glutamine transaminase with improved enzymatic activity, wherein the amino acid sequence is shown as SEQ ID NO.1. According to the invention, amino acid at the N end of MTG (Microbial Transglutaminase) maturase is subjected to deficiency and saturation mutation by taking the efficient expression of MTG in colibacillus as an improvement platform, and a mutant strain with good enzymatic property is obtained. The enzyme activity is improved by 1.85 times, and the thermal stability is improved by 2.7 times. The modified enzyme is more suitable for industrial applications, the production cost can be lowered, and the production efficiency can be improved.

Description

technical field [0001] The invention relates to a glutamine transaminase, in particular to a glutamine transaminase with improved enzyme activity and thermal stability. Background technique [0002] The biological function of microbial transglutaminase (protein-glutamic acid-glutaminase, Microbial Transglutaminase, EC2.3.2.13 referred to as MTG) is to directly change the protein itself and the cells and tissues to which the protein is attached. The structural and functional properties of protein enhance the nutritional value of protein. Therefore, MTG has broad application prospects in food, textile, biopharmaceutical and other fields. However, due to some defects of MTG itself, such as specific enzyme activity, thermal stability and other factors, the scope of application of MTG is limited. Therefore, based on the obtained expression platform of MTG in Escherichia coli, amino acid deletion and saturation mutation were used to carry out molecular transformation of MTG, in ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/10C12N15/70C12R1/55
Inventor 陈坚陈康康刘松张东旭马建龙堵国成
Owner JIANGNAN UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com