Nucleotide sequence analysis and heterologous expression of the Erysipelothrix rhusiopathiae dnaJ gene

Abstract DNA sequence analysis of chromosomal DNA from the Gram-positive facultative intracellular pathogen, Erysipelothrix rhusiopathiae has identified a dnaJ heat shock gene homolog. A 1109-bp open reading frame encoding dnaJ is located immediately 3' to the E. rhusiopathiae dnaK gene. The deduced DnaJ amino acid sequence exhibits the modular structure of other members of the DnaJ protein class including a glycine-rich region and the repeating consensus sequence CXXCXGXGX. Heterologous expression of the dnaJ sequence in Escherichia coli resulted in accumulation of a unique 38.9-kDa protein with an isoelectric point of 8.0. Deletion analysis of the dnaJ gene was used to confirm that the overproduced protein was encoded by the dnaJ sequence.     

The nucleotide sequence of the Escherichia coli K12 dnaJ+ gene. A gene that encodes a heat shock protein

The Escherichia coli dnaJ gene product is required for bacteriophage lambda DNA replication at all temperatures. It is also essential for bacterial viability in at least some conditions, since mutations in it result in temperature-sensitive bacterial growth. We have previously cloned the dnaJ gene and shown that its product migrates as a Mr 37,000 polypeptide under denaturing conditions. Here we present the primary DNA sequence of the dnaJ gene. It codes for a processed basic protein (63 basic and 51 acidic amino acids) composed of 375 amino acids totaling Mr 40,973. The predicted NH2-terminal amino acid sequence, overall amino acid composition, and isoelectric point agree well with those of the purified protein. We present evidence that the rate of expression of the dnaJ protein is increased by heat shock under the control of the htpR (rpoH) gene product.     

Characterization of ApuB, an extracellular type II amylopullulanase from Bifidobacterium breve UCC2003

The apuB gene of Bifidobacterium breve UCC2003 was shown to encode an extracellular amylopullulanase. ApuB is composed of a distinct N-terminally located alpha-amylase-containing domain which hydrolyzes alpha-1,4-glucosidic linkages in starch and related polysaccharides and a C-terminally located pullulanase-containing domain which hydrolyzes alpha-1,6 linkages in pullulan, allowing the classification of this enzyme as a bifunctional class II pullulanase. A knockout mutation of the apuB gene in B. breve UCC2003 rendered the resulting mutant incapable of growth in medium containing starch, amylopectin, glycogen, or pullulan as the sole carbon and energy source, confirming the crucial physiological role of this gene in starch metabolism.