Preparation method and application of halophilic archaea extracellular protease truncation

Abstract

The invention belongs to the field of gene engineering, and relates to a preparation method and application of a halophilic archaea extracellular protease truncated body. The method comprises the following steps: transforming a prokaryotic host by using a recombinant plasmid pET28a-hlyA, carrying out induced expression to obtain halophilic archaea protease, carrying out purification and renaturation on the halophilic archaea protease, and incubating to obtain the high-purity halophilic archaea extracellular protease truncation HlyA CTE of which the amino acid sequence is that a mature enzyme sequence of HlyA lacks a CTE structural domain (SEQ ID No.1). The HlyA CTE has excellent enzymatic characteristics, and particularly has good stability in high-temperature, low-salt, high-salt, alkaline and other environments; meanwhile, the HlyA CTE can be used for catalyzing hydrolysis of protein substrates, including catalysis of macromolecular protein substrates represented by azocasein, is specifically applied to fish sauce fermentation to promote hydrolysis of fish protein, and provides a novel enzyme preparation for protein degradation in high-salt, high-temperature and other environments.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and in particular relates to a preparation method and application of a highly stable halophilic archaea extracellular protease truncated body. Background technique [0002] Protease is a general term for a class of enzymes that hydrolyze protein peptide chains. It is widely used in food, biomedicine, leather, textile and detergent industries, but its application is limited under extreme conditions. Halophilic archaea usually live in high-salt environments such as salt lakes, saline-alkali land, drying salt fields, and pickled foods. Therefore, halophilic archaea are a good source of universal protease, which can be used in special industrial conditions such as high salt and high temperature. [0003] At present, the published halophilic archaeal extracellular proteases are all serine proteases, which belong to the subtilase S8A family. The discovered proteases are all formed intracellularly i...

AI Technical Summary

Problems solved by technology

[0004] The protease produced by halophilic archaea will be depolymerized or inactivated under low-salt conditions, so there are certain limitations in its separation and purification

At present, the purification methods of protease include bacitracin affinity chromatography, gel filtration chromatography and other methods. The proteins obtained by these methods generally contain both the target band and the impurity band, and further purification is required, and the stability is not good

Images