Plasmid location, cloning, and sequence analysis of the gene encoding a 27.3-kilodalton cytolytic protein from Bacillus thuringiensis subsp. morrisoni (PG-14)

A gene encoding the 27.3 kilodalton cytolytic protein toxin in the mosquitocidal isolate (PG-14) of Bacillus thuringiensis subsp. morrisoni (BTM) was cloned and sequenced, and compared with the homologous gene in B. thuringiensis subsp. israelensis (BTI). The BTM gene was determined to be located on a 140 kb plasmid by use of a synthetic 20-base nucleotide probe derived from the sequence of the BTI gene. A 9.4-kb Hind III plasmid fragment containing the BTM cytolytic protein gene was cloned into the plasmid vector pUC12, and subsequently subcloned and sequenced. Comparison of the nucleotide sequence of the BTM gene with that of the homologous BTI gene revealed only a single base difference; the base at position 310 in BTM is guanine, whereas in BTI it is cytosine. This single base change results in the occurrence of alanine rather than proline at amino acid residue 82 in BTM. Analysis of the secondary structure and hydropathic profile of the BTM protein indicates that alanine at this position increases both the propensity to form an α-helical structure and the hydrophobicity in the vicinity of this residue. Thus, the BTM toxin is potentially more cytolytic than the homologous protein of BTI.