The plasmid pBMBt1 from Bacillus thuringiensis subsp. darmstadiensis (INTA Mo14-4) replicates by the rolling-circle mechanism and encodes a novel insecticidal crystal protein-like gene.
This work describes a novel rolling-circle replicating (RCR) plasmid pBMBt1 from Bacillus thuringiensis subsp. darmstadiensis (INTA Mo14-4) encoding an insecticidal crystal protein-like gene. pBMBt1 (6700 bp) contains three ORFs and their putative transcription initiation sites and Shine-Dalgarno sequences were localized. ORF1 encodes a 34.6 kDa protein which showed identity with the protein CryC53 from B. thuringiensis subsp. cameroun (24.6%), the Cry15Aa insecticidal crystal protein from B. thuringiensis subsp. thompsoni (21.9%) and the Mtx3 protein from Bacillus sphaericus (27.8%). The ORF2 (52.3 kDa) showed a 74% identity with the Mob protein coded by pUIBI-1 from B. thuringiensis subsp. entomocidus and 64% identity with the Mob protein of pBMY1 from Bacillus mycoides; both Mob proteins belong to the pMV158 superfamily. To evaluate the Mob protein, the plasmid pHTMob14-4 was constructed. This plasmid shows transfer frequencies of 9.1x10(-6) in B. thuringiensis subsp. israelensis (4Q7Gm(R)). The ORF3 (23.6 kDa) gene product is homologous to the Rep protein from the plasmid pBMYdx of B. mycoides (37.6%). A putative double-strand origin with significant homology to that of B. thuringiensis plasmids, and an ssoA-type single-strand origin were also identified. Detection of single-stranded pBMBt1 DNA replicating intermediaries suggests that replication occurs via the rolling-circle mechanism.
Loeza-Lara
Pedro D
PD
Centro Multidisciplinario de Estudios en Biotecnología-Facultad de Medicina Veterinaria y Zootecnia, Universidad Michoacana de San Nicolás de Hidalgo, Apdo. Postal 53, Administración Chapultepec, C.P. 58262 Morelia, Michoacán, Mexico.
Benintende
Graciela
G
Cozzi
Jorge
J
Ochoa-Zarzosa
Alejandra
A
Baizabal-Aguirre
Victor M
VM
Valdez-Alarcón
Juan J
JJ
López-Meza
Joel E
JE
D000595:Amino Acid Sequence; D001413:Bacillus thuringiensis / Q000235:genetics*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics; D001427:Bacterial Toxins / Q000737:chemistry / Q000235:genetics; D001483:Base Sequence; D017124:Conserved Sequence; D004261:DNA Replication; D004269:DNA, Bacterial; D004270:DNA, Circular; D004731:Endotoxins / Q000737:chemistry / Q000235:genetics; D006460:Hemolysin Proteins; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D010957:Plasmids / Q000235:genetics*; D017386:Sequence Homology, Amino Acid
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004269:DNA, Bacterial; D004270:DNA, Circular; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis