US 11,932,881 B2Granted

Heparin skeleton synthase and its mutants and application

AssigneesShan Dong University, Bloomage Biotechnology Corporation Limited

InventorsJuzheng Sheng, Xueping Guo, Jianqun Deng, Fengshan Wang, Zhen Lu, Ranran Du, Liu Sun, Yuanjun Sun

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Abstract

A heparin skeleton synthase originates from Neisseria animaloris, with an amino acid sequence as shown in SEQ ID NO.2 and a nucleotide sequence of the coding gene as shown in SEQ ID NO.1. Its recombinant expression level is 6.8 times that of the existing heparin skeleton synthase KfiA from Escherichia coli K5, and total enzyme activity per fermentation liquor is 5.22 times that of the heparin skeleton synthase KfiA. The heparin skeleton synthase mutants obtained through site-directed mutagenesis of the sites No. 16, No. 25, No. 30, No. 111, No. 165, and No. 172 in the amino acid sequence of the said heparin skeleton synthase all have high expression levels.

Claims7

Dependent claims

A mutant heparin skeleton synthase (NaGlcNAc-T) is obtained from a wild-type NaGlcNAc-T, wherein the mutant NaGlcNAc-T is selected from the group consisting of a mutant heparin skeleton synthase having C16L mutation and comprises the amino acid sequence of SEQ ID NO:

4, a mutant heparin skeleton synthase having N25P mutation and comprises the amino acid sequence of SEQ ID NO:
6, a mutant heparin skeleton synthase having I30L mutation and comprises the amino acid sequence of SEQ ID NO:
8, a mutant heparin skeleton synthase having I111S mutation and comprises the amino acid sequence of SEQ ID NO:
10, a mutant heparin skeleton synthase having S165K mutation and comprises the amino acid sequence of SEQ ID NO:
12, a mutant heparin skeleton synthase having S172A mutation and comprises the amino acid sequence of SEQ ID NO:
14.

3 more elements

Dependencies are inferred from the claim text.

Facts

Application: 17/646,773Filed 3 Jan 2022
Priority: CN 202110192424.220 Feb 2021
Published: US 2022/0267746 A125 Aug 2022
Granted: 19 Mar 2024
Term ends: Not estimatedThe source records don’t support an estimate.

Classification10 codes

C12N 9/1051
Glycosyltransferases: hexosyltransferases
C12N 15/102
Processes for the isolation, preparation or purification of DNA or RNA: mutagenizing nucleic acids
C12N 15/70
Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression: vectors or expression systems…
C12P 19/46
Preparation of O-glycosides, e.g. glucosides: having an oxygen atom of the saccharide radical bound to a cyclohexyl radical, e.g. kasugamycin