Biomaterial Database

Cloning and characterization of the dnaK heat shock operon of the marine bacterium Vibrio harveyi. 1998

G Klein, and M Zmijewski, and J Krzewska, and M Czeczatka, and B Lipińska

Department of Biochemistry, University of Gdańsk, Poland.

We cloned the DNA region of the Vibrio harveyi chromosome containing the heat shock genes dnaK and dnaJ and sequenced them. These genes are arranged in the chromosome in the order dnaK-dnaJ, as in other proteobacteria of the alpha and gamma subdivisions. The dnaK gene is 1923 nucleotides in length and codes for a protein of 640 amino acid residues, with a predicted molecular mass of 69,076 Da and 81.2% similarity to the DnaK protein of Escherichia coli. The V. harveyi dnaJ gene has a coding sequence of 1158 nucleotides. The predicted DnaJ protein contains 385 amino acids, its calculated molecular mass is 41,619 Da and it has 74.7% similarity to the DnaJ protein of E. coli. Northern hybridization experiments with RNA from V. harveyi cells and a DNA probe carrying both the dnaK and dnaJ genes showed a single, heat-inducible transcript, indicating that these genes form an operon. Primer extension analysis revealed five heat-inducible transcriptional start sites upstream of the dnaK gene, two of which (T1 and T4) are preceded by sequences typical of the E. coli heat shock promoters recognized by the sigma 32 (sigma32) factor. Location of these promoters is highly similar to that of the E. coli dnaK promoters. No transcriptional start sites were detected upstream of the dnaJ gene. The V. harveyi dnaKJ operon cloned in a plasmid in E. coli cells was transcribed in a sigma32 dependent manner and the size of the transcript, the kinetics of transcription, and the transcriptional start sites were as in V. harveyi cells. This indicates a high conservation of the transcriptional heat shock regulatory elements between E. coli and V. harveyi, both belonging to the gamma subdivision of proteobacteria. We tested the ability of the cloned dnaKJ genes to complement E. coli dnaK and dnaJ mutants and found that V. harveyi DnaJ restored a thermoresistant phenotype to dnaJ mutants and enabled lambda phage to grow in the mutant cells. V. harveyi DnaK did not suppress the thermosensitivity of dnaK mutants but complemented the dnaK deletion mutant with respect to growth of lambda phage. V. harveyi DnaK, in contrast to DnaJ, failed to modulate the heat shock response in E. coli. Our results suggest that the DnaK chaperone may be more species specific than the DnaJ chaperone.

UI	MeSH Term	Description	Entries
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D009876	Operon	In bacteria, a group of metabolically related genes, with a common promoter, whose transcription into a single polycistronic MESSENGER RNA is under the control of an OPERATOR REGION.	Operons
D003001	Cloning, Molecular	The insertion of recombinant DNA molecules from prokaryotic and/or eukaryotic sources into a replicating vehicle, such as a plasmid or virus vector, and the introduction of the resultant hybrid molecules into recipient cells without altering the viability of those cells.	Molecular Cloning
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D005798	Genes, Bacterial	The functional hereditary units of BACTERIA.	Bacterial Gene,Bacterial Genes,Gene, Bacterial
D005816	Genetic Complementation Test	A test used to determine whether or not complementation (compensation in the form of dominance) will occur in a cell with a given mutant phenotype when another mutant genome, encoding the same mutant phenotype, is introduced into that cell.	Allelism Test,Cis Test,Cis-Trans Test,Complementation Test,Trans Test,Allelism Tests,Cis Tests,Cis Trans Test,Cis-Trans Tests,Complementation Test, Genetic,Complementation Tests,Complementation Tests, Genetic,Genetic Complementation Tests,Trans Tests
D006360	Heat-Shock Proteins	Proteins which are synthesized in eukaryotic organisms and bacteria in response to hyperthermia and other environmental stresses. They increase thermal tolerance and perform functions essential to cell survival under these conditions.	Stress Protein,Stress Proteins,Heat-Shock Protein,Heat Shock Protein,Heat Shock Proteins,Protein, Stress
D000595	Amino Acid Sequence	The order of amino acids as they occur in a polypeptide chain. This is referred to as the primary structure of proteins. It is of fundamental importance in determining PROTEIN CONFORMATION.	Protein Structure, Primary,Amino Acid Sequences,Sequence, Amino Acid,Sequences, Amino Acid,Primary Protein Structure,Primary Protein Structures,Protein Structures, Primary,Structure, Primary Protein,Structures, Primary Protein
D001483	Base Sequence	The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.	DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA
D012329	RNA, Bacterial	Ribonucleic acid in bacteria having regulatory and catalytic roles as well as involvement in protein synthesis.	Bacterial RNA