TY - JOUR
T1 - Parasporin 1Ac2, a novel cytotoxic crystal protein isolated from Bacillus thuringiensis B0462 strain
AU - Kuroda, Shouta
AU - Begum, Anowara
AU - Saga, Mizue
AU - Hirao, Akina
AU - Mizuki, Eiichi
AU - Sakai, Hiroshi
AU - Hayakawa, Tohru
N1 - Funding Information:
This work was supported, in part, by Grant-in-Aid for Scientific Research (20380036, 23658048 and 24380034).
PY - 2013/5
Y1 - 2013/5
N2 - Two novel parasporin (PS) genes were cloned from Bacillus thuringiensis B0462 strain. One was 100 % identical even in nucleotide sequence level with that of parasporin-1Aa (PS1Aa1) from B. thuringiensis A1190 strain. The other (PS1Ac2) showed significant homology (99 % identity) to that of PS1Ac1 from B. thuringiensis 87-29 strain. The 15 kDa (S113-R250) and 60 kDa (I251-S777) fragments consisting of an active form of PS1Ac2 were expressed as His-tag fusion. Upon purification under denaturing condition and refolding, the recombinant polypeptides were applied to cancer cells to analyze their cytotoxicities. 3-(4,5-Dimethyl-2-thiazoyl)-2,5-diphenyl- 2H-tetrazolium bromide assay revealed that either of 15 or 60 kDa polypeptide exhibited no cytotoxicity to HeLa cells, but they became cytotoxic upon mixed together. Our results suggested that PS1Ac2 was responsible for the cytotoxicity of B. thuringiensis B0462 strain, and that the formation of hetero-dimer of 15 and 60 kDa polypeptide was required for their cytotoxicity.
AB - Two novel parasporin (PS) genes were cloned from Bacillus thuringiensis B0462 strain. One was 100 % identical even in nucleotide sequence level with that of parasporin-1Aa (PS1Aa1) from B. thuringiensis A1190 strain. The other (PS1Ac2) showed significant homology (99 % identity) to that of PS1Ac1 from B. thuringiensis 87-29 strain. The 15 kDa (S113-R250) and 60 kDa (I251-S777) fragments consisting of an active form of PS1Ac2 were expressed as His-tag fusion. Upon purification under denaturing condition and refolding, the recombinant polypeptides were applied to cancer cells to analyze their cytotoxicities. 3-(4,5-Dimethyl-2-thiazoyl)-2,5-diphenyl- 2H-tetrazolium bromide assay revealed that either of 15 or 60 kDa polypeptide exhibited no cytotoxicity to HeLa cells, but they became cytotoxic upon mixed together. Our results suggested that PS1Ac2 was responsible for the cytotoxicity of B. thuringiensis B0462 strain, and that the formation of hetero-dimer of 15 and 60 kDa polypeptide was required for their cytotoxicity.
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U2 - 10.1007/s00284-013-0301-1
DO - 10.1007/s00284-013-0301-1
M3 - Article
C2 - 23306354
AN - SCOPUS:84876682088
SN - 0343-8651
VL - 66
SP - 475
EP - 480
JO - Current Microbiology
JF - Current Microbiology
IS - 5
ER -
