Mutant of alpha-L-rhamnosidase from aspergillus terreus CCF 3059 and application thereof

Abstract

The invention provides a mutant of alpha-L-rhamnosidase from aspergillus terreus CCF 3059 and application thereof. The mutant comprises a gene D594Q shown in SEQ ID NO:2, a gene D594R shown in SEQ ID NO:3, a gene D594C shown in SEQ ID NO:4, a gene G827K shown in SEQ ID NO:5, a gene G827M shown in SEQ ID NO:6 and a gene G828A shown in SEQ ID NO:7. The mutant has the beneficial effects that the optimum temperatures of a mutant enzyme MRha-D594Q and a proenzyme MRha are 65 DEG C, but compared with the proenzyme MRha, the mutant enzyme MRha-D594Q still maintains higher enzymatic activity at 70 DEG C and 75 DEG C; the heat stability of the mutant enzyme MRha-D594Q can be further improved by adding sorbitol and is improved by 7.8 times in the half-life period at 70 DEG C.

Description

Technical field [0001] The invention belongs to the field of molecular biology and enzyme engineering, and specifically relates to an Aspergillus terreus CCF 3059 α-L-rhamnosidase mutant and its application. Background technique [0002] α-L-rhamnosidase (EC 3.2.1.40) is an important glycoside hydrolase, which can specifically hydrolyze many natural glycoside compounds containing non-reducing α-L-rhamnose at the end, such as reed Ding, naringin, hesperidin, stevioside, myricetin, etc. The source of α-L-rhamnosidase is relatively wide, and it is widely distributed in plants, bacteria, fungi, and animal livers in nature. The classification of glycoside hydrolase in the CAZy (http: / / www.cazy.org / ) database is based on the similarity of amino acid sequences. α-L-rhamnosidase is included in three glycoside hydrolase families (GH ), respectively GH13, GH78 and GH106. [0003] α-L-rhamnosidase has a wide range of applications in the food industry. As we all know, many fruits contain b...

AI Technical Summary

Problems solved by technology

However, there are still obstacles and technical bottlenecks in the large-scale application of α-L-rhamnosidase in industry: (1) the activity of existing strains to produce α-L-rhamnosidase is not high enough, which directly leads to the use cost higher

(2) The optimal reaction temperature of most reported α-L-rhamnosidases is between 40 and 60°C, and the thermal stability is poor under high temperature conditions, which is not conducive to the dissolution of the substrate and the purification of the product

Although the thermal stability of the enzyme can be improved by the above techniques, there are no reports utilizing these techniques, especially two or more methods to improve the stability of α-L-rhamnosidase

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