Nucleotide sequence of a novel delta-endotoxin gene cryIg of Bacillus thuringiensis ssp. galleriae.

A gene cryIg coding for entomocidal protein delta-endotoxin of Bacillus thuringiensis ssp. galleriae str. 11-67 named CryIG has been cloned and sequenced (EMBL accession number X58120). The deduced amino acid sequence that contains 1156 amino acid residues shows only 28% of identical residues, when compared with other delta-endotoxins of the CryI family. The extent of identity is substantially higher for some regions of the sequence ('conserved blocks'), that presumably bear important structural or functional properties. This implies that CryIG delta-endotoxin follows the same type of polypeptide chain folding as other CryI proteins, whereas peculiarities of primary structure help to explain its unique specificity.

Molecular cloning and primary structure of Thermoactinomyces vulgaris carboxypeptidase T. A metalloenzyme endowed with dual substrate specificity.

A gene coding for an extracellular Zn-carboxypeptidase of Thermoactinomyces vulgaris has been cloned and sequenced (EMBL X56901). This enzyme named carboxypeptidase T reveals simultaneously both types of substrate specificity characteristic of mammalian carboxypeptidases A and B. The carboxypeptidase T gene is primarily expressed in E. coli as a non-active preproenzyme with an additional 98 amino acid residues at the N-terminus. Primary structure alignment of mature carboxypeptidase T and mammalian metallocarboxypeptidases demonstrated 25-30% overall identity but a full preservation of presumed catalytically important residues. These observations imply a basic uniformity of the general catalytic mechanism for enzymes of that class produced by evolutionarily remote organisms.

[Multiple genes of delta-endotoxins from Bacillus thuringiensis subspecies galleriae]. / Mnozhestvennye geny delta-éndotoksinov Bacillus thuringiensis podvida galleriae.

Cloning and expression in E. coli delta-endotoxin genes cryIAb7, cryIG and, cryIX from Bacillus thuringiensis ssp galleriae has been performed. Restriction mapping and partial sequencing have shown the identity of the 3'-terminal parts of cryIG and cryIX, although their 5'-terminal halves corresponding to the toxin are unique. Sequencing 5'-flanking region of cryIX revealed no translation initiation site, which indicates a nonfunctional state of the gene. The clusterized locating of the cryIG and cryIG genes has been shown. The absence of a promoter-like structure in the 5'-flanking region of cryIG suggests transcription of the gene as a bicistronic mRNA with cryIX. The extremely high homology of the cryIG and cryIX 3'-terminal parts (2 kB long) suggests a recombination act in the gene origin.

[Creation of a hybrid protein gene based on Bacillus thuringiensis delta endotoxins CryIIIA and CrIA(a) and expression of its derivatives in Escherichia coli]. / Sozdanie gena gibridnogo belka na osnove delta-éndotoksinov Bacillus thuringiensis CryIIIA i CryIA(a) i ékspressiia ego proizvodnykh v Escherichia coli.

The 5'-terminal fragment (containing 1-565th codons) of Bacillus thuringiensis var. tenebrionis gene for the Coleoptera-specific delta-endotoxin CryIIIA was cloned. This sequence was extended with either only a homologous fragment of CryIA(a) or that one together with in-frame NPTII or GUS coding sequences. The obtained gene derivatives were expressed in E. coli. The analysis of hybrid polypeptides confirmed the enzymatic activities of bifunctional proteins and showed toxic properties of "insectotoxin-NPTII" against Colorado potato beetle (Leptinotarsa decemlineata).