C-type lectin as well as preparation method and application thereof

A technology of lectin and derivatives, which is applied in the field of C-type lectin and its preparation and application, and can solve the problems of the structure, preparation and biological function of C-type lectin that have not been seen before.

Active Publication Date: 2014-06-04
SHENYANG PHARMA UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] So far, there is no research on the structure, preparation and biological function of C-type lectin in Lepidoptera (Lepidoptera) insects.

Method used

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  • C-type lectin as well as preparation method and application thereof
  • C-type lectin as well as preparation method and application thereof
  • C-type lectin as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0069] Separation and purification of natural C-type lectin

[0070] 1. method 1

[0071] The pre-cooled saturated ammonium sulfate was stirred and slowly added to the hemolymph for fractional precipitation of ammonium sulfate. Take 30-50% of the precipitated fraction to dissolve and dilute with citric acid-phosphate buffer system, and use this as the initial loading solution to load the sample on the Q-Sepharose anion chromatography column for gradient elution. The components containing the target protein were loaded on the SP-Sepharose cationic chromatography column for gradient elution, and each washed and eluted fraction was tracked and monitored by Western blot with C-type lectin antibody. Finally, the gel filtration chromatography column Sepharose CL-6B was used to perform double separation and purification of the above components by gel filtration and affinity chromatography, so as to obtain pure C-type lectin with natural activity.

[0072] A single component was ...

Embodiment 2

[0080] C-type lectin structure analysis and its gene sequence analysis

[0081] According to the techniques, methods and means of conventional protein chemistry and molecular biology, the structure analysis of C-type lectins was carried out. The specific methods and means are implemented according to the contents listed in the second content of the invention.

[0082] Obtain the primary structure-amino acid sequence of natural C-type lectin (also known as mature peptide chain, referred to as C-type lectin in this patent), as well as the gene encoding natural C-type lectin and its sequence (such as figure 2 shown).

[0083] Using bioinformatics technology, methods, etc., the structure of the C-type lectin of the present invention was analyzed by homologous comparison, and the results showed that: the C-type lectin of the present invention belongs to the C-type lectin superfamily member, and the sequence and its homologous The comparison of the ten insect-derived lectins with...

Embodiment 3

[0085]Using prokaryotic expression system to obtain recombinant C-type lectin and its derivatives, analogs, and active fragments

[0086] This example lists and describes the construction strategy and basic method for expressing the C-type lectin and its derivatives, analogs, and active fragment genes of the present invention in a prokaryotic expression system.

[0087] The expression vector, expression host cell and expression strategy of the prokaryotic expression system are the conventional and general expression vector, expression host cell and expression strategy of genetic engineering expression.

[0088] This implementation is to enable those skilled in the art to understand the present invention more comprehensively, but not to limit the scope of the special claims of the present invention in any way.

[0089] For the separation and purification method of the expression product, the method, principle, strategy, etc. of Example 1 were adopted.

[0090] 1. Construction ...

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Abstract

The invention relates to the technical field of biological medicines, and in particular relates to an obtainment method of a C-type lectin and a derivative, an analogue or an active fragment thereof as well as applications of the C-type lectin and the derivative, the analogue or the active fragment thereof in mode detection of microorganisms and related molecules, induction of insects to generate antimicrobial peptides, preparation of an antibody of the C-type lectin and the derivative, the analogue or the active fragment thereof and the like. The amino acid sequence of the C-type lectin is shown as the figure 2-(A). The sequence of the derivative, the analogue or the active fragment of the C-type lectin is shown as the figure 3. The C-type lectin with an electrophoresis purity and even a HPLC (High Performance Liquid Chromatography) purity is obtained by performing ion-exchange chromatography or hydrophobic chromatography or affinity chromatography or gel filtration or salting or ultrafiltration separation and purification on the raw materials of the C-type lectin, or treating the raw materials of the C-type lectin through the combination of two or multiple methods among the ion-exchange chromatography, the hydrophobic chromatography, the affinity chromatography, the gel filtration, the salting or the ultrafiltration separation and purification and according to the recombination of the separation and purification methods.

Description

technical field [0001] The present invention relates to the field of biomedical technology, and relates to the structure of C-type lectin and its obtaining method and application. Specifically, the present invention relates to the structure and production method of C-type lectin and its derivatives, analogs, and active fragments , and its applications in the detection of microorganisms and their related molecular patterns, inducing insects to produce antimicrobial peptides, and preparing C-type lectins and their derivatives, analogs, and active fragment antibodies. Background technique [0002] Lectins are a class of non-enzyme and non-antibody proteins that can specifically recognize sugar structures and bind non-covalently and reversibly to them, and their activity depends on Ca 2+ , known as C-type lectin (C-type lectin). Due to the wide variety, they constitute a huge C-type lectin superfamily (CLSF). Most of its members are integrated transmembrane proteins, and there...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/435C12N15/12C07K1/18C07K1/22C07K1/20C07K1/16C07K1/30C07K1/34C07K1/36C12N15/63C12N15/70C12N15/75C12N15/81C12N15/85C07K16/18
CPCC07K14/7056C07K16/2851
Inventor 张嵘张景海吴春福
Owner SHENYANG PHARMA UNIVERSITY
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