Catechol dioxygenase mutant, recombinant vector as well as preparation method and application of catechol dioxygenase mutant
A catechol dioxygenase and dioxygenase technology, applied in the biological field, can solve the problems of limited industrial application and low thermal stability, and achieve the effect of large application potential and high thermal stability
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0060] This embodiment provides a mutant catechol dioxygenase, the mutant catechol dioxygenase is as follows (a1) or (a2):
[0061] (a1) A polypeptide having the same function as the polypeptide composed of the amino acid sequence shown in SEQ ID No.2 obtained by deleting, replacing or adding one or more amino acids from the amino acid sequence shown in SEQ ID No.2;
[0062] (a2) A polypeptide having at least 90% homology to the polypeptide consisting of the amino acid sequence shown in SEQ ID No.2.
[0063] Specifically, in a preferred embodiment, the mutant catechol dioxygenase is mutated on the amino acid sequence shown in SEQ ID No.2, and the mutation site is selected from A95V, L148S, R193Q, L248D one or a combination of more than one.
[0064] In a further preferred embodiment, the mutation site is A95V, L148S, R193Q, L248D, A95V-L148S, L148S-R193Q, A95V-L248D, R193Q-L248D, A95V-R193Q, L148S-L248D, A95V-L148S-R193Q, A95V-L148S-L248D, A95V-R193Q-L248D, L148S-R193Q-L248D...
Embodiment 2
[0067] This embodiment provides a gene encoding the mutant catechol dioxygenase in embodiment 1, the gene is as follows (b1) or (b2):
[0068] (b1) A polynucleotide obtained by deleting, replacing or adding one or more bases from the nucleotide sequence shown in SEQ ID No.1;
[0069] (b2) A polynucleotide having at least 90% homology to the polynucleotide consisting of the nucleotide sequence shown in SEQ ID No.1.
[0070] Specifically, in a preferred embodiment, the nucleotide sequence encoding the mutant catechol dioxygenase whose mutation site is A95V is shown in SEQ ID No.28; the mutant type whose encoding mutation site is L148S The nucleotide sequence of catechol dioxygenase is shown in SEQ ID No.29; the nucleotide sequence of the mutant catechol dioxygenase encoding mutation site R193Q is shown in SEQ ID No.30 Shown; The nucleotide sequence of the mutant type catechol dioxygenase encoding mutation site is L248D as shown in SEQ ID No.31; The encoding mutation site is the...
Embodiment 3
[0072] This embodiment provides a recombinant vector comprising the gene of Embodiment 1.
[0073]The preparation method of the recombinant vector comprises the following steps: using a pair of mutation amplification primers to perform PCR amplification on the first vector to obtain the recombinant vector, wherein the pair of mutation amplification primers contains a mutation site of mutant catechol dioxygenase The nucleotide sequence corresponding to the point, the first vector contains the gene encoding the amino acid sequence shown in SEQ ID No.2.
[0074] Wherein, the mutation amplification primer pair includes: the amplification primer pair of amplification mutation site A95V as shown in SEQ ID No.20 and SEQ ID No.21, as shown in SEQ ID No.22 and SEQ ID No.23 The amplification primer pair of the amplification mutation site L148S, the amplification primer pair of the amplification mutation site R193Q shown in SEQ ID No.24 and SEQ ID No.25, such as SEQ ID No.26 and SEQ ID N...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More