Expression and purification of the BmK M1 neurotoxin from the scorpion Buthus martensii Karsch.
The gene encoding a neurotoxin (BmK M1) from the scorpion Buthus martensii Karsch was expressed in Saccharomyces cerevisiae at a high level with the alcohol dehydrogenase promoter. SDS-PAGE of the culture confirmed expression and showed secretion into medium from yeast. Recombinant BmK M1 was purified rapidly and efficiently by ion exchange and gel filtration chromatography to homogeneity, produced a single band on tricine-SDS-PAGE, and processed the homologous N-terminus. Amino acid analysis and N-terminal sequencing demonstrated that the recombinant toxin was processed correctly from the alpha-mating factor leader sequence and was chemically identical to the native form. The expressed recombinant BmK M1 was toxic for mice, which indicated that it was biologically active. Quantitative estimation showed that recombinant BmK M1 had an LD(50) similar to that of the native toxin.
Shao
F
F
Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101, People's Republic of China.
Xiong
Y M
YM
Zhu
R H
RH
Ling
M H
MH
Chi
C W
CW
Wang
D C
DC
D000818:Animals; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors; D007928:Lethal Dose 50; D051379:Mice; D009498:Neurotoxins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D012604:Scorpion Venoms / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D020539:Sequence Analysis, Protein
D009498:Neurotoxins; D011993:Recombinant Fusion Proteins; D012604:Scorpion Venoms