Expression and Characterization of Two Kinds of Recombinant Snake Neurotoxins
10.3321/j.issn:1000-3061.2000.03.004
- VernacularTitle:蛇神经毒素的表达和鉴定
- Author:
You-Cun QIAN
1
;
Yan SHEN
;
Chun-Yang FAN
;
Tai-Shan HU
;
Sheng-Li YANG
;
Yi GONG
Author Information
1. Shanghai Research Center of Biotechnology, The Chinese Academy of Sciences
- Keywords:
Neurotoxin;
fusion expression;
genetic engineering
- From:
Chinese Journal of Biotechnology
2000;16(3):312-315
- CountryChina
- Language:Chinese
-
Abstract:
The cDNA encoding the precursor of cobrotoxin was cloned from the venom gland of the Chinese continental cobra ( Naja naja atra ) by RT-PCR. Its deduced amino acid sequence analysis showed that the mature protein was identical to that identified from the Taiwan cobra ( Naja naja atra ) by protein sequencing technique. The cDNA encoding the mature protein was then subcloned into the expression vector pMAL-P2. The gene of CMl 1, which was formed by ligation of the fragments of the synthetic oligonucleotides, was also cloned into the expression vector pMAL-P2. After induction of IPTG, both of the neurotoxins were overexpressed as soluble fusion proteins which were confirmed by SDS-PAGE and western blotting. The expressed fusion proteins was purified by sepharose 6B-amylose affinity chromatography and DEAE-sepharose FF chromatography. Both of the recombinant proteins achieved after digestion by factor Xa showed the in vivo toxicity.
