Genomic DNA sequences and functional expression, purification of BmalphaTX14 neurotoxin from scorpion Buthus martensii Karsch.
- Author:
Meng LÜ
1
;
Kun WANG
;
Zhi-Jian CAO
;
Da-He JIANG
;
Xin MAO
;
Wen-Xin LI
Author Information
1. State Key Lab of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China.
- Publication Type:Journal Article
- MeSH:
Amino Acid Sequence;
Animals;
Base Sequence;
Cloning, Molecular;
Molecular Sequence Data;
Pichia;
genetics;
metabolism;
Recombinant Proteins;
biosynthesis;
genetics;
isolation & purification;
Scorpion Venoms;
biosynthesis;
genetics;
Scorpions;
genetics;
Sequence Analysis
- From:
Chinese Journal of Biotechnology
2005;21(6):853-857
- CountryChina
- Language:Chinese
-
Abstract:
Based on the full-length cDNA of BmalphaTX14 from Chinese scorpion Buthus martensii Karsch (BmK), gene of the mature peptide of BmalphaTX14 was cloned into the yeast expression vector pPIC9K. After transforming, screening and inducing, tricine-SDS-PAGE and Western blot proved that rBmalphaTX14 protein was expressed in the medium for up to 84 hours, getting nearly 120 mg/L. Recombinant BmalphaTX14 was purified rapidly and efficiently through Ni-NTA-agarose, polyethylene glycol precipitation and gel filtration chromatography. The purified rBmalphaTX14 proved to have the anti-insect activity by toxicity assay. Meanwhile, genomic gene of BmalphaTX14 was cloned and sequenced by PCR method, sequence analysis of this gene showed that BmalphaTX14 had an intron of 408 base pairs located at the signal peptide encoding region, which was similar with the characteristic of other alpha-type sodium ion-channel toxin. Considering both the genomic organization and the peptide function, BmaTX14 proved to be a membership belonging to alpha-type sodium ion-channel toxin.
