The Intermediate Domain Defines Broad Nucleotide Selectivity for Protein Folding in Chlamydophila GroEL1

Okuda, Hiroshi; Sakuhana, Chihaya; Yamamoto, Risa; Mizukami, Yuko; Kawai, Rika; Sumita, Yusuke; Koga, Motoki; Shirai, Mutsunori; Matsuda, Kazuhiko

doi:10.1074/jbc.m710057200

Cited by 4 publications

(3 citation statements)

References 35 publications

(30 reference statements)

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“…1 and Table 1). A prior study with ChGroEL from C. pneumoniae reported ATP hydrolysis at a rate 50% of that of EcGroEL (22). ChGroES inhibited the ATPase activity of ChGroEL by 50%, the same inhibitory effect seen for EcGroES with EcGroEL (24)(25)(26)(27)(28).…”

Section: Resultsmentioning

confidence: 75%

“…Sequence identity between ChGroEL and EcGroEL is high (64%), while that between ChGroEL2 or ChGroEL3 and EcGroEL is low (23% or 32%, respectively). Previous research found that ChGroEL-ChGroES assists in the folding of proteins, including malate dehydrogenase (MDH) (22), and can replace EcGroEL-EcGroES to support the growth of E. coli (23), suggesting a conventional folding chaperone function for ChGroEL. By contrast, ChGroEL2 and ChGroEL3 cannot substitute for EcGroEL (23), suggesting that neither paralog duplicates the conventional folding function of a chaperonin.…”

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confidence: 99%

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Biochemical and Genetic Analysis of the Chlamydia GroEL Chaperonins

et al. 2017

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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 chaperonin, EcGroEL, and its cochaperonin, EcGroES, assist in the folding of proteins via an ATP-dependent mechanism. In addition to the presence of and homologs, paralogs are found in many bacteria, including pathogens, and have evolved poorly understood species-specific functions. spp., which are obligate intracellular bacteria, have reduced genomes that nonetheless contain three genes, (), , and We hypothesized that ChGroEL is the bona fide chaperonin and that the paralogs perform novel -specific functions. To test our hypothesis, we investigated the biochemical properties of ChGroEL and its cochaperonin, ChGroES, and queried the essentiality of the three genes through targeted mutagenesis in 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 , 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. 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 and, which might indicate a divergent evolution of GroEL during the transition from a free-living organism to an obligate intracellular lifestyle.

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Section: Resultsmentioning

confidence: 75%

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confidence: 99%

Biochemical and Genetic Analysis of the Chlamydia GroEL Chaperonins

et al. 2017

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“…GroEL ATPase assays ( 60 ), MDH ( 61 ) and LDH ( 62 ) refolding assays, and the proteinase K protection assay ( 17 ) were performed following previously described protocols. A detailed description of such assays is available in the Supplementary Materials.…”

Section: Methodsmentioning

confidence: 99%

Engineering a nanopore with co-chaperonin function

Meervelt

Tsai

et al. 2015

Sci. Adv.

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Distinct features of protein folding by the GroEL system from a psychrophilic bacterium, Colwellia psychrerythraea 34H

et al. 2012

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We investigated the protein folding mechanism of the GroEL system of a psychrophilic bacterium, Colwellia psychrerythraea 34H. The amount of mRNA of the groESL operon of C. psychrerythraea was increased about 6-fold after a temperature upshift from 8 to 18 °C for 30 min, suggesting that this temperature causes heat stress in this bacterium. A σ(32)-type promoter was found upstream of the groESL, suggesting that the C. psychrerythraea groESL is regulated by the σ(32) system, like the groESL in E. coli. The maximum ATPase and CTPase activities of CpGroEL were observed at 45 and 35 °C, respectively, which are much higher than the growth temperatures of C. psychrerythraea. We found that the refolding activity of the CpGroEL system in the presence of ATP is lower than that in the presence of CTP. This suggests that ATP is not the optimum energy source of the CpGroEL system. Analyses for the interaction of CpGroEL-CpGroES revealed that CTP could weaken this interaction, resulting in effective refolding function of the CpGroEL system. From these findings, we consider that the CpGroEL system possesses an energy-saving mechanism for avoiding excess consumption of ATP to ensure growth in a low-temperature environment.

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The Intermediate Domain Defines Broad Nucleotide Selectivity for Protein Folding in Chlamydophila GroEL1

Cited by 4 publications

References 35 publications

Biochemical and Genetic Analysis of the Chlamydia GroEL Chaperonins

Biochemical and Genetic Analysis of the Chlamydia GroEL Chaperonins

Engineering a nanopore with co-chaperonin function

Distinct features of protein folding by the GroEL system from a psychrophilic bacterium, Colwellia psychrerythraea 34H

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