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CN-122012435-A - Glycosyltransferase mutant and application thereof in production of salidroside

CN122012435ACN 122012435 ACN122012435 ACN 122012435ACN-122012435-A

Abstract

The invention discloses a glycosyltransferase mutant and application thereof in production of salidroside, and relates to the technical field of biology. The glycosyltransferase mutant is any one of A1) to A2), wherein A1) is protein obtained by carrying out amino acid mutation on at least one of the 41 st, 114 th, 272 th and 340 th positions of the amino acid sequence of wild type glycosyltransferase, A2) is fusion protein obtained by connecting a tag with the N end or/and the C end of the protein shown in A1), and the amino acid sequence of the wild type glycosyltransferase is shown as SEQ ID NO. 1. It can efficiently catalyze tyrosol to generate salidroside.

Inventors

  • LIU JIANZHONG
  • ZHAO SIBO

Assignees

  • 中山大学

Dates

Publication Date
20260512
Application Date
20260122

Claims (10)

  1. 1. A glycosyltransferase mutant, characterized in that the glycosyltransferase mutant is any one of A1) to A2): A1 A protein obtained by mutating at least one of the 41 st, 114 th, 272 th and 340 th positions of the amino acid sequence of the wild type glycosyltransferase; A2 Fusion proteins obtained by ligating a tag to the N-terminal or/and the C-terminal of the protein represented by A1); the amino acid sequence of the wild glycosyltransferase is shown as SEQ ID NO. 1.
  2. 2. The glycosyltransferase mutant according to claim 1, wherein the glycosyltransferase mutant is a protein obtained by amino acid mutation at position 41 of the amino acid sequence of a wild-type glycosyltransferase, a protein obtained by amino acid mutation at position 114 of the amino acid sequence of a wild-type glycosyltransferase, a protein obtained by amino acid mutation at position 272 of the amino acid sequence of a wild-type glycosyltransferase, a protein obtained by amino acid mutation at position 340 of the amino acid sequence of a wild-type glycosyltransferase or a protein obtained by amino acid mutation at positions 41, 114, 272 and 340 of the amino acid sequence of a wild-type glycosyltransferase, preferably wherein the amino acid mutation at position 41 is Y41H, and/or wherein the amino acid mutation at position 114 is G114A, and/or wherein the amino acid mutation at position 340 is S272T, and/or wherein the amino acid mutation at position 340 is A340T.
  3. 3. A biomaterial characterized in that the biomaterial is any one of B1) to B4): B1 A nucleic acid molecule encoding the glycosyltransferase mutant of claim 1 or 2; B2 An expression cassette comprising the nucleic acid molecule of B1); b3 A recombinant vector comprising the nucleic acid molecule of B1) or a recombinant vector comprising the expression cassette of B2); B4 A recombinant biological cell comprising B1) said nucleic acid molecule, B2) said expression cassette or B3) said recombinant vector.
  4. 4. The biomaterial of claim 3, wherein the nucleic acid molecule of B1) is any one of the following: the nucleotide sequence of the nucleic acid molecule B1-1) is shown as SEQ ID NO. 11, SEQ ID NO. 12, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15; B1-2) has more than 75% identity to the nucleotide sequence set forth in B1-1) and encodes a cDNA molecule or a DNA molecule of the glycosyltransferase mutant set forth in the examples of the first aspect; B1-3) hybridizes under stringent conditions to a nucleotide sequence defined in any one of B1-1) -B1-2) and encodes a cDNA molecule or DNA molecule of the glycosyltransferase mutant described in the examples of the first aspect.
  5. 5. Use of the biomaterial of claim 3 or 4 in any one of C1) to C3): C1 Preparing the glycosyltransferase mutant of claim 1 or 2; C2 Preparing salidroside; C3 Preparing a product for generating salidroside.
  6. 6. The use of claim 5, wherein the product comprises at least one of a reagent, a kit.
  7. 7. Use of a glycosyltransferase mutant according to claim 1 or 2 for the preparation of a product comprising or producing salidroside.
  8. 8. A method for preparing salidroside, which is characterized by comprising the following steps: Catalyzing the production of salidroside from tyrosol with the glycosyltransferase mutant of claim 1 or 2 or recombinant cells expressing the glycosyltransferase mutant.
  9. 9. The method of claim 8, wherein the catalytic temperature is 25 ℃ to 35 ℃.
  10. 10. An enzyme preparation comprising the glycosyltransferase mutant of claim 1 or 2.

Description

Glycosyltransferase mutant and application thereof in production of salidroside Technical Field The invention relates to the technical field of biology, in particular to a glycosyltransferase mutant and application thereof in production of salidroside. Background Salidroside is a glycosylation product of tyrosol which is a main active substance of rhodiola rosea. The salidroside has the functions of resisting anoxia, cold, fatigue, microwave radiation, virus, tumor and the like, has the effects of enhancing attention, improving working efficiency, delaying organism aging, preventing senile diseases and the like, has very important application value in the aspects of military medicine, aerospace medicine, sports medicine, health care medicine and the like, and is an environment-adaptive medicine with great development prospect, and attention is paid in recent years. In the related art, salidroside is generally obtained by taking rhodiola plants as raw materials through an extraction method, and has the problems of high extraction cost, low yield and the like. And the salidroside is synthesized by constructing engineering bacteria and fermenting, so that the problem of low synthesis efficiency exists, and industrialization is difficult to realize. Therefore, there is a need to provide a method that can efficiently synthesize salidroside. Disclosure of Invention The present invention aims to solve at least one of the technical problems existing in the prior art. To this end, the invention proposes a glycosyltransferase mutant. The invention also provides biological materials related to the glycosyltransferase mutants. The invention also provides application of the biological material. The invention also provides application of the glycosyltransferase mutant. The invention also provides a method for preparing the salidroside. The invention also provides an enzyme preparation comprising the glycosyltransferase mutant. A glycosyltransferase mutant according to an embodiment of the first aspect of the present invention, which is any one of A1) to A2): A1 A protein obtained by mutating at least one of the 41 st, 114 th, 272 th and 340 th positions of the amino acid sequence of the wild type glycosyltransferase; A2 Fusion proteins obtained by ligating a tag to the N-terminal or/and the C-terminal of the protein represented by A1); the amino acid sequence of the wild glycosyltransferase is shown as SEQ ID NO. 1. The glycosyltransferase mutant provided by the embodiment of the invention has at least the following beneficial effects: the glycosyltransferase mutant can efficiently catalyze tyrosol to generate salidroside, and has higher catalytic activity than wild glycosyltransferase. The capability of the microorganism expressing glycosyltransferase mutant for converting tyrosol into salidroside is obviously improved, and the method has good application prospect. According to some embodiments of the invention, the glycosyltransferase mutant is a protein obtained by amino acid mutation at position 41 of the amino acid sequence of the wild-type glycosyltransferase. According to some embodiments of the invention, the glycosyltransferase mutant is a protein obtained by amino acid mutation at position 114 of the amino acid sequence of the wild-type glycosyltransferase. According to some embodiments of the invention, the glycosyltransferase mutant is a protein obtained by amino acid mutation at position 272 of the amino acid sequence of a wild-type glycosyltransferase. According to some embodiments of the invention, the glycosyltransferase mutant is a protein obtained by amino acid mutation at position 340 of the amino acid sequence of the wild-type glycosyltransferase. According to some embodiments of the invention, the glycosyltransferase mutant is a protein obtained by mutating amino acids at positions 41, 114, 272 and 340 of the amino acid sequence of the wild-type glycosyltransferase. According to some embodiments of the invention, the amino acid at position 41 is mutated to Y41H. According to some embodiments of the invention, the amino acid at position 114 is mutated to G114A. According to some embodiments of the invention, the amino acid mutation at position 272 is S272T. According to some embodiments of the invention, the amino acid at position 340 is mutated to a340T. According to some embodiments of the invention, the tag comprises at least one of a tag that facilitates solubilization, purification and detection of the glycosyltransferase mutant. It will be appreciated that the glycosyltransferase mutants of the invention may comprise one or more tags, and that the plurality of tags may comprise a combination of a plurality of identical tags, or may be a combination of a plurality of different tags. For example, labels that facilitate solubilization of the glycosyltransferase mutant include, but are not limited to, nus labels or maltose binding protein labels, labels that facilitate purification of the glycosyltr