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Strong Secretory Signal Peptide Enhancing Small Peptide Motif and Its Application

A signal peptide, secretory technology, applied in the field of protein engineering and genetic engineering, to achieve the effect of enhancing the secretion significantly, the effect is obvious, and the secretion function is enhanced

Active Publication Date: 2017-11-28
NANJING TECH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] Although the extracellular secretion expression strategy has significant advantages over other expression methods, there are still many limitations in the widespread use of this strategy in E. coli systems

Method used

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  • Strong Secretory Signal Peptide Enhancing Small Peptide Motif and Its Application
  • Strong Secretory Signal Peptide Enhancing Small Peptide Motif and Its Application
  • Strong Secretory Signal Peptide Enhancing Small Peptide Motif and Its Application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Example 1. Application of Small Peptide Enhancement Motif in Fructosidase Secretion and Expression

[0062] First, the source of the fructosidase Fru6 gene described in this example is: Arthrobacter arilaitensis NJEM01 strain, which is the inventor's prior Chinese patent application CN102732456A, and the strain's deposit number is: CCTCCNO: M2012155.

[0063] All primers involved in this example were synthesized by Yingjun Company, as shown in Table 1. The following primer numbers are uniformly represented by "P" followed by a sequence number. For example, primer No. 45 in Table 2 has its code P45, and its number in the sequence listing is SEQ ID NO: 45.

[0064] Table 1 Nucleotide sequences of primers and synthetic sequences required in Example 1

[0065]

[0066]

[0067] Note: The underlined part in the table shows the restriction enzyme cleavage site.

[0068] (1) Fructosidase FRU6 gene acquisition: Using the genome of Arthrobacter arilaitensis NJEM01 as a t...

Embodiment 2

[0090] Example 2 Application of Small Peptide Enhancement Motif in Secretion and Expression of Glucanase

[0091] (1) Construction of expression vector for the expression vector of glucanase BGL by adding small peptides before different signal peptides

[0092] Similar to Example 1, in this example, the fructosidase in Example 1 was replaced with a glucanase gene.

[0093] Among them, the source of the sequence of glucanase BGL: Bacillus subtilis subsp. subtilis 6051-HGW, GenBank serial number: CP003329.1, range: 4011849 to 4012490.

[0094] The primers P79 and P80 were used to amplify the gene fragment of glucanase BGL by PCR reaction, and the T vector pMD-T-BGL was prepared. Overlapping PCR primers P81-P84 or P85-P88 fused glucanase BGL and signal peptide ompA or pelB, respectively, wherein P84 and P88 both carry EcoRI enzyme cleavage sites. P89-P96 upstream primers are used in combination with downstream primers P84 or P88, and different enhancer motifs can be designed to...

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Abstract

The invention belongs to the field of protein engineering and genetic engineering, and relates to a class of strongly secreted signal peptide enhancing small peptide modules and applications thereof. The strong secretory signal peptide enhancing small peptide motif of the present invention has an amino acid sequence of the following general formula: M(αXβYγ / αYβXγ)n, wherein X represents an acidic amino acid; Y represents a basic amino acid; α is 0-2 β represents 0-2 neutral amino acids; γ represents 1-10 neutral amino acids; n is 1-3. The application of the strong secretory signal peptide enhancing small peptide motif of the present invention is to use the strong secretory signal peptide enhancing small peptide motif of the present invention to construct a carrier that enhances the secretory ability of ordinary signal peptides, so as to improve It secretes a method of expressing a foreign protein.

Description

[0001] This application is a divisional application for an invention patent application. The application date of the original application is May 3, 2013, the application number is 201310162455.9, and the name of the invention is "Strong Secretory Signal Peptide Enhancement Small Peptide Motif and Its Application". technical field [0002] The invention belongs to the fields of protein engineering and genetic engineering, and relates to a type of strong secretory signal peptide enhancing small peptide motif and its application. Background technique [0003] The protein secretion system is not only the premise of protein sorting, localization and realization of physiological functions, but also an important means of genetic engineering and protein engineering technology development. Various prokaryotic and eukaryotic expression systems have been developed for the production of recombinant proteins. Escherichia coli expression system has attracted much attention due to its cle...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/06C12N15/11C12N15/70C12N15/67C12N1/21
Inventor 何冰芳陈文华米兰吴珊珊朱芸陈珊珊
Owner NANJING TECH UNIV
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