Cloning and characterization of a mosquito larvicidal toxin produced during vegetative stage of Bacillus sphaericus 2297.
The mosquitocidal toxin 1 (mtx1) gene from genomic DNA of B. sphaericus strain 2297 was cloned and expressed in E. coli. DNA sequencing analysis of the cloned gene revealed a single open reading frame encoding an 870-amino acid polypeptide. Expression level of the full-length gene in E. coli was very low even though strong promoter was used or the gene was expressed as a fusion protein. Expression level was highly improved after the putative leader sequence was deleted, and the truncated gene was expressed as a fusion protein with glutathione S-transferase (GST-tMtx1). E. coli cells expressing GST-tMtx1 was highly toxic to Culex quinquefasciatus larvae and showed lower toxicity against Anopheles dirus and Aedes aegypti larvae. Enterobacter amnigenus An11, a mosquito larval gut colonizable bacteria, transformed with the cloned gene exhibited mosquito larvicidal activity. Result suggested that there is a potential to develop this protein to be used as an alternative mosquito control agent.
National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, 113 Paholyothin Road, Klong 1, Klong Luang, Pathumthani 12120, Thailand. boonhiang@biotec.or.th
Boonhiang
B
Promdonkoy
Patcharee
P
Promdonkoy
Sutipa
S
Tanapongpipat
Plearnpis
P
Luxananil
Namchai
N
Chewawiwat
Mongkon
M
Audtho
Sakol
S
Panyim
D000818:Animals; D001407:Bacillus / Q000235:genetics* / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics* / Q000494:pharmacology*; D001427:Bacterial Toxins / Q000235:genetics* / Q000494:pharmacology*; D003001:Cloning, Molecular; D009033:Culicidae / Q000187:drug effects*; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification; D004754:Enterobacter / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D007306:Insecticides / Q000494:pharmacology; D007814:Larva / Q000187:drug effects; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals / Q000235:genetics / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004269:DNA, Bacterial; D007306:Insecticides; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C046134:binB protein, Bacillus sphaericus; D005982:Glutathione Transferase