Nucleotide sequence and transcriptional analysis of activator-regulator proteins for beta-lactamase in Streptomyces cacaoi.
The nucleotide sequence of the 2.7-kb DNA fragment upstream of the structural gene of beta-lactamase in Streptomyces cacaoi was determined. Computer-aided "FRAME" analysis revealed four possible open reading frames (ORFs), three in one direction and one in the opposite direction. One of them (ORF1, BlaA) encoded an activator-regulator protein whose deduced amino acid sequence was similar to that of other activator-regulator proteins in bacteria. Insertion of an 8-bp BamHI linker into the BlaA region decreased the beta-lactamase activity sharply, from 50 U to 1 U/ml. This protein (BlaA) was found to bind to the nucleotide sequence between the bla (beta-lactamase structural gene) and blaA genes. Another ORF (ORF2, BlaB) in the same orientation had a couple of amino acid sequences similar to that of pBR322 beta-lactamase. However, insertion of the 8-bp BamHI linker indicated that this ORF was functional as an activator-regulator but not as a beta-lactamase. Therefore, there were two activator-regulator proteins in the upstream region of the structural gene of the beta-lactamase. Nuclease S1 mapping predicted that transcription for the activator proteins commenced at the translational initiation codon or within a few nucleotides from the translational start site. Transcription was in the opposite direction to that of the beta-lactamase structural gene.
Urabe
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Department of Biochemistry, Meiji College of Pharmacy, Tokyo, Japan.
Ogawara
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D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001483:Base Sequence; D002874:Chromosome Mapping; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000235:genetics*; D005798:Genes, Bacterial / Q000235:genetics*; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D016364:Reading Frames; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D013302:Streptomyces / Q000235:genetics*; D014158:Transcription, Genetic; D001618:beta-Lactamases / Q000096:biosynthesis / Q000235:genetics*
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C074160:BlaA protein, Bacteria; D001618:beta-Lactamases