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Bullfrog lectin gene, encoded lectin and uses thereof

A technology of lectin and bullfrog, applied in the field of genetic engineering, can solve problems such as toxicity and immunogenic side effects, and achieve the effects of inhibiting bacterial growth, convenient artificial synthesis and simple structure

Inactive Publication Date: 2011-09-14
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For decades, researchers have been looking for suitable lectin drug delivery carrier molecules, but most of the known lectins have a molecular weight greater than 10,000, and large molecular weight lectins may cause toxicity and Immunogenicity and other serious side effects, small molecular weight lectins will overcome these defects and serve as ideal drug delivery carrier molecules

Method used

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  • Bullfrog lectin gene, encoded lectin and uses thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Bullfrog lectin gene clone:

[0041] I. Extraction of Total RNA from Bullfrog Skin

[0042] A. The live bullfrog was cleaned with double-distilled water, and the medullary cavity of the bullfrog was destroyed with a sterilized needle. 50-100 mg of bullfrog skin tissue was put into a dry-baked mortar, and 1 mL of TRIZOL Reagent (product of Invitrogen Company) was added to quickly Grind in an ice water bath.

[0043] B. After fully mixing, transfer to a 1.5mL centrifuge tube (DEPC tube) and place at 15-30°C for 5min, add an equal volume of chloroform, vortex mix for 15s, and centrifuge at 12000×g for 15min at 4°C.

[0044] C. Add 500 μL of isopropanol to the supernatant, place at 15-30°C for 5 minutes, vortex for 15 seconds to mix well, and centrifuge at 12,000×g for 10 minutes at 4°C. Discard the supernatant, add 1 mL of 75% ethanol to rinse the precipitate, centrifuge at 7500×g for 5 min, and repeat once. Dry in an ultra-clean workbench for 90 seconds, dissolve with ...

Embodiment 2

[0089] Bullfrog Lectin

[0090] I. Sample preparation method: the amino acid sequence of bullfrog lectin was deduced according to the gene encoding bullfrog lectin, and its complete sequence was synthesized with an automatic polypeptide synthesizer APEX396. Desalted and purified by HPLC reverse phase C18 column chromatography.

[0091] II. Molecular weight was determined by electrospray ionization (ESI) with an emission voltage of 4.5Kv.

[0092] III. The purity of the purified bullfrog lectin was identified by high performance liquid chromatography HPLC (flow rate 1.0mL / min; mobile phase acetonitrile+water+TFA0.01%, gradient elution; chromatographic column C18, detection wavelength 220nm), molecular weight determination The isoelectric point was determined by fast atom bombardment mass spectrometry and isoelectric focusing electrophoresis, and the amino acid sequence structure was determined by an automatic amino acid sequencer.

[0093] Bullfrog lectin is a single-chain po...

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Abstract

The invention relates to a bullfrog agglutinin gene, agglutinin encoded by the same, and application in pharmacy, belonging to the technical field of gene engineering. The bullfrog agglutinin gene is screened from a constructed bullfrog skin cDNA library with molecule weight of 71.8 Dalton and isoelectric point of 8.75; and a primary structure of polypeptide complete sequence is Phe Leu Thr Phe Pro Gly Met Thr Phe Gly Lys Leu Leu-AMIDATION; C-end is alpha-amidation modification. The gene encoding the bullfrog agglutinin contains 330 bits of nucleotides, wherein encoded mature peptides are nucleotides from 145 to 183 bit. Artificially synthesized bullfrog agglutinin has obvious effects of agglutinating rabbit erythrocyte and inhibiting bacterium growth; due to the advantages of small molecular weight and simple synthesis, the bullfrog agglutinin obtained by the gene can be used in antibacterial feed additive, drug development and a functional molecule which plays a leading role in vector targeting transportation of drug.

Description

technical field [0001] The invention relates to a bullfrog lectin gene, the encoded lectin and its application, and belongs to the technical field of genetic engineering. Background technique [0002] Lectin is a protein that can be synthesized and secreted by both animal cells and plant cells and can bind sugar. It plays an important role in cell recognition and adhesion reactions, mainly promoting intercellular adhesion. The biggest feature of lectin is that it can recognize complex carbohydrate structures in glycoproteins and glycolipids, especially in cell membranes, that is, sugar groups on the surface of cell membranes. Lectin has more than one sugar-binding site, so it can participate in cell recognition and adhesion, and connect different cells. Lectins have multiple biological activities in invertebrate blood, can selectively agglutinate various cells, have specific agglutination effects on tumor cells, etc., and are one of the important humoral factors of immune d...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/12C07K7/08A23K1/16A61K47/42A61K38/10A61P31/04A61P7/04A23K20/153A23K50/80
Inventor 韩文瑜赵瑞利韩俊友雷连成孙长江冯新
Owner JILIN UNIV