Transcriptional and translational start sites for the Bacillus thuringiensis crystal protein gene

Abstract

The nucleotide sequence of the promoter region and part of the coding region of the crystal protein gene from Bacillus thuringiensis var. kurstaki HD-1-Dipel has been determined by analysis of a recombinant plasmid from Escherichia coli. The start points for transcription of the gene in B. thuringiensis and in the E. coli strain carrying the recombinant plasmid were located by S1 nuclease mapping. Two adjacent start sites were identified using RNAs synthesized during sporulation of B. thuringiensis: transcription was initiated from one site early in sporulation and from the other site in the middle of sporulation. A good correlation was found between the appearance of the crystal protein gene-specific RNA and the production of the protein, indicating that the gene is primarily under transcriptional control during sporulation. Parallel studies with the recombinant strain of E. coli revealed the presence of only a single species of gene-specific RNA, regardless of the growth phase of the cells; the crystal protein was produced at all stages of growth. The sequence for eight amino acids at the NH2 terminus of the crystal protein was determined and the corresponding coding sequence was located in the DNA sequence. A potential ribosome binding site of 11 nucleotides was found, located three nucleotides upstream from the initiator ATG codon. The deduced sequence for the first 333 amino acids of the crystal protein is presented.