Study on construct and expression of synthetic genes encoding spider dragline silk in Escherichia coli.
- Author:
Min LI
1
;
Wen-Xian ZHANG
;
Zhi-Hua HUANG
;
Jian-Kun HUANG
Author Information
1. College of Biological Engineering, Fujian Normal University, Fuzhou 350007, China. MLI2@sina.com
- Publication Type:Journal Article
- MeSH:
Amino Acid Sequence;
Amino Acids;
analysis;
Base Sequence;
Escherichia coli;
genetics;
Fibroins;
Molecular Sequence Data;
Molecular Weight;
Proteins;
analysis;
genetics;
isolation & purification;
Recombinant Proteins;
analysis;
isolation & purification
- From:
Chinese Journal of Biotechnology
2002;18(3):331-334
- CountryChina
- Language:Chinese
-
Abstract:
Dragline spider silk produced from Nephilia clavipes major ampullate is a natural fibrous protein with specific mechanical properties such as high tensile strength and elasticity. Synthetic gene monomer encoding recombinant spider silk protein, based on the known repetitive protein sequence and partial cDNA sequence of dragline silk, was constructed and expressed. DNA monomer sequences were multimerized to encode high molecular weight synthetic spider silks using a "head-to-tail" construction strategy. Multimer was cloned into pET30a(+), a prokaryotic high potency expression vector, and induced with IPTG. The protein from 8-unit repeat was produced in Escherichia coli at levels up to 20 mg/L. The protein was easily purified with high recovery by using an metal ion affinity chromatography and purity was over 90%. The results of SDS-PAGE and Western blot suggested that the mass of the expression product was about 37 kD. This value and amino acid analysis were consistent with those of theoretic calculation.
