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Differentiation of rickettsiae by groEL gene analysis.

ABSTRACT: The nucleotide sequences (534 to 546 bp) of the groEL gene, which encodes the 60-kDa heat shock protein GroEL, from 15 rickettsial strains were determined and compared. In the phylogenetic tree created by the unweighted pair group method with arithmetic averages and the neighbor-joining method, rickettsial strains could be distinguished from Ehrlichia strains. Five spotted fever group strains, four typhus group strains, and six scrub typhus group (STG) strains were differentiated as distinct entities. Unlike gltA and ompA gene analyses, differentiation between members of the genus Rickettsia and the STG rickettsiae by groEL gene analysis was possible. In comparison with 16S rRNA gene analysis, the groEL gene has a higher degree of divergence among the rickettsiae. We therefore successfully developed rapid differentiation methods, PCR-restriction fragment length polymorphism analysis and a species-specific PCR, based on the groEL gene sequences. Four Korean isolates were identified by these methods and groEL gene analysis. The results suggest that the groEL gene is useful for the identification and characterization of rickettsiae.

SUBMITTER: Lee JH

PROVIDER: S-EPMC165385 | biostudies-literature | 2003 Jul

REPOSITORIES: biostudies-literature

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Differentiation of rickettsiae by groEL gene analysis.

Lee Jung-Hee JH Park Hyo-Soon HS Jang Won-Jong WJ Koh Seong-Eun SE Kim Jong-Moon JM Shim Soo-Kyoung SK Park Mi-Yeoun MY Kim Yoon-Won YW Kim Bum-Joon BJ Kook Yoon-Hoh YH Park Kyung-Hee KH Lee Seung-Hyun SH

Journal of clinical microbiology 20030701 7

The nucleotide sequences (534 to 546 bp) of the groEL gene, which encodes the 60-kDa heat shock protein GroEL, from 15 rickettsial strains were determined and compared. In the phylogenetic tree created by the unweighted pair group method with arithmetic averages and the neighbor-joining method, rickettsial strains could be distinguished from Ehrlichia strains. Five spotted fever group strains, four typhus group strains, and six scrub typhus group (STG) strains were differentiated as distinct ent ...[more]

PMID: 12843026

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Project description:Chaperonins are essential for cellular growth under normal and stressful conditions and consequently represent one of the most conserved and ancient protein classes. The paradigm Escherichia coli chaperonin, EcGroEL, and its cochaperonin, EcGroES, assist in the folding of proteins via an ATP-dependent mechanism. In addition to the presence of groEL and groES homologs, groEL paralogs are found in many bacteria, including pathogens, and have evolved poorly understood species-specific functions. Chlamydia spp., which are obligate intracellular bacteria, have reduced genomes that nonetheless contain three groEL genes, Chlamydia groEL (ChgroEL), ChgroEL2, and ChgroEL3 We hypothesized that ChGroEL is the bona fide chaperonin and that the paralogs perform novel Chlamydia-specific functions. To test our hypothesis, we investigated the biochemical properties of ChGroEL and its cochaperonin, ChGroES, and queried the in vivo essentiality of the three ChgroEL genes through targeted mutagenesis in Chlamydia trachomatis ChGroEL hydrolyzed ATP at a rate 25% of that of EcGroEL and bound with high affinity to ChGroES, and the ChGroEL-ChGroES complex could refold malate dehydrogenase (MDH). The chlamydial ChGroEL was selective for its cognate cochaperonin, ChGroES, while EcGroEL could function with both EcGroES and ChGroES. A P35T ChGroES mutant (ChGroESP35T) reduced ChGroEL-ChGroES interactions and MDH folding activities but was tolerated by EcGroEL. Both ChGroEL-ChGroES and EcGroEL-ChGroESP35T could complement an EcGroEL-EcGroES mutant. Finally, we successfully inactivated both paralogs but not ChgroEL, leading to minor growth defects in cell culture that were not exacerbated by heat stress. Collectively, our results support novel functions for the paralogs and solidify ChGroEL as a bona fide chaperonin that is biochemically distinct from EcGroEL.IMPORTANCEChlamydia is an important cause of human diseases, including pneumonia, sexually transmitted infections, and trachoma. The chlamydial chaperonin ChGroEL and chaperonin paralog ChGroEL2 have been associated with survival under stress conditions, and ChGroEL is linked with immunopathology elicited by chlamydial infections. However, their exact roles in bacterial survival and disease remain unclear. Our results further substantiate the hypotheses that ChGroEL is the primary chlamydial chaperonin and that the paralogs play specialized roles during infection. Furthermore, ChGroEL and the mitochondrial GroEL only functioned with their cochaperonin, in contrast to the promiscuous nature of GroEL from E. coli and Helicobacter pylori, which might indicate a divergent evolution of GroEL during the transition from a free-living organism to an obligate intracellular lifestyle.

Dataset Information

Differentiation of rickettsiae by groEL gene analysis.

Publications

Differentiation of rickettsiae by groEL gene analysis.

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