Acid protease mutant as well as coding gene, preparation method and application thereof

Abstract

The invention belongs to the technical field of gene engineering, and particularly relates to an acid protease mutant as well as a coding gene, a preparation method and application thereof. The amino acid sequence of the acid protease mutant is as shown in SEQ ID NO. 2. The acid protease mutant provided by the invention has the advantages of high enzyme activity, strong pH and temperature adaptability, good high temperature resistance and the like, can react in a wide pH value (pH 2.0-6.0) and temperature range (30-70 DEG C), is beneficial for improving the flexibility of a production process, particularly can adapt to industrial production at a relatively high temperature and can meet a relatively long-time high-temperature catalytic environment, and the acid protease mutant has a wide application prospect. The enzyme prepared by the method is high in enzyme activity, not easy to inactivate in a high-temperature environment, can maintain 70% or more of enzyme activity for 1 h at 50 DEG C, can maintain 55% or more of enzyme activity for 1 h at 55 DEG C, and shows good market prospects and industrial application values in the fields of wine brewing, feed processing, leather softening and the like.

Claims

1. An acidic protease mutant, characterized in that, The amino acid sequence is shown in SEQ ID NO.

2.

2. A nucleic acid molecule, characterized in that, Includes the gene sequence encoding the acidic protease mutant as described in claim 1.

3. The nucleic acid molecule according to claim 2, characterized in that, The gene sequence of the acidic protease mutant is shown in SEQ ID NO.

3.

4. The nucleic acid molecule according to claim 2, characterized in that, The nucleic acid molecule also includes gene sequences encoding signal peptides, promoters, terminators, resistance markers, and / or replication initiation sites.

5. The nucleic acid molecule according to claim 4, characterized in that, The nucleotide sequences of the signal peptide gene, promoter gene, terminator gene, replication initiation site gene, and resistance marker gene are shown in SEQ ID NO.4-8, respectively.

6. An expression carrier, characterized in that, The expression vector comprises the nucleic acid molecule as described in any one of claims 3-5.

7. A recombinant bacterial strain, characterized in that, The recombinant strain comprises the expression vector as described in claim 6.

8. The recombinant strain according to claim 7, characterized in that, The recombinant strain originated from Trichoderma reesei QM 9414.

9. The method for preparing the acidic protease mutant according to claim 1, characterized in that, Includes the following steps: Cultivate the recombinant strain as described in any one of claims 7-8, centrifuge the fermentation culture broth, filter the supernatant, and collect the filtrate to obtain the crude enzyme solution containing the acidic protease mutant.

10. The use of the acidic protease mutant as described in claim 1 in brewing, feed processing, or leather softening.

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