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Distinct properties of hexameric but functionally conserved Mycobacterium tuberculosis transcription-repair coupling factor.

ABSTRACT: Transcription coupled nucleotide excision repair (TC-NER) is involved in correcting UV-induced damage and other road-blocks encountered in the transcribed strand. Mutation frequency decline (Mfd) is a transcription repair coupling factor, involved in repair of template strand during transcription. Mfd from M. tuberculosis (MtbMfd) is 1234 amino-acids long harboring characteristic modules for different activities. Mtbmfd complemented Escherichia coli mfd (Ecomfd) deficient strain, enhanced survival of UV irradiated cells and increased the road-block repression in vivo. The protein exhibited ATPase activity, which was stimulated ?1.5-fold in the presence of DNA. While the C-terminal domain (CTD) comprising amino acids 630 to 1234 showed ?2-fold elevated ATPase activity than MtbMfd, the N-terminal domain (NTD) containing the first 433 amino acid residues was able to bind ATP but deficient in hydrolysis. Overexpression of NTD of MtbMfd led to growth defect and hypersensitivity to UV light. Deletion of 184 amino acids from the C-terminal end of MtbMfd (Mfd?C) increased the ATPase activity by ?10-fold and correspondingly exhibited efficient translocation along DNA as compared to the MtbMfd and CTD. Surprisingly, MtbMfd was found to be distributed in monomer and hexamer forms both in vivo and in vitro and the monomer showed increased susceptibility to proteases compared to the hexamer. Mfd?C, on the other hand, was predominantly monomeric in solution implicating the extreme C-terminal region in oligomerization of the protein. Thus, although the MtbMfd resembles EcoMfd in many of its reaction characteristics, some of its hitherto unknown distinct properties hint at its species specific role in mycobacteria during transcription-coupled repair.

SUBMITTER: Prabha S

PROVIDER: S-EPMC3084762 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature

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Distinct properties of hexameric but functionally conserved Mycobacterium tuberculosis transcription-repair coupling factor.

Prabha Swayam S Rao Desirazu N DN Nagaraja Valakunja V

PloS one 20110429 4

Transcription coupled nucleotide excision repair (TC-NER) is involved in correcting UV-induced damage and other road-blocks encountered in the transcribed strand. Mutation frequency decline (Mfd) is a transcription repair coupling factor, involved in repair of template strand during transcription. Mfd from M. tuberculosis (MtbMfd) is 1234 amino-acids long harboring characteristic modules for different activities. Mtbmfd complemented Escherichia coli mfd (Ecomfd) deficient strain, enhanced surviv ...[more]

PMID: 21559463

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Project description:UnlabelledWhile SecA is the ATPase component of the major bacterial secretory (Sec) system, mycobacteria and some Gram-positive pathogens have a second paralog, SecA2. In bacteria with two SecA paralogs, each SecA is functionally distinct, and they cannot compensate for one another. Compared to SecA1, SecA2 exports a distinct and smaller set of substrates, some of which have roles in virulence. In the mycobacterial system, some SecA2-dependent substrates lack a signal peptide, while others contain a signal peptide but possess features in the mature protein that necessitate a role for SecA2 in their export. It is unclear how SecA2 functions in protein export, and one open question is whether SecA2 works with the canonical SecYEG channel to export proteins. In this study, we report the structure of Mycobacterium tuberculosis SecA2 (MtbSecA2), which is the first structure of any SecA2 protein. A high level of structural similarity is observed between SecA2 and SecA1. The major structural difference is the absence of the helical wing domain, which is likely to play a role in how MtbSecA2 recognizes its unique substrates. Importantly, structural features critical to the interaction between SecA1 and SecYEG are preserved in SecA2. Furthermore, suppressor mutations of a dominant-negative secA2 mutant map to the surface of SecA2 and help identify functional regions of SecA2 that may promote interactions with SecYEG or the translocating polypeptide substrate. These results support a model in which the mycobacterial SecA2 works with SecYEG.ImportanceSecA2 is a paralog of SecA1, which is the ATPase of the canonical bacterial Sec secretion system. SecA2 has a nonredundant function with SecA1, and SecA2 exports a distinct and smaller set of substrates than SecA1. This work reports the crystal structure of SecA2 of Mycobacterium tuberculosis (the first SecA2 structure reported for any organism). Many of the structural features of SecA1 are conserved in the SecA2 structure, including putative contacts with the SecYEG channel. Several structural differences are also identified that could relate to the unique function and selectivity of SecA2. Suppressor mutations of a secA2 mutant map to the surface of SecA2 and help identify functional regions of SecA2 that may promote interactions with SecYEG.

Dataset Information

Distinct properties of hexameric but functionally conserved Mycobacterium tuberculosis transcription-repair coupling factor.

Publications

Distinct properties of hexameric but functionally conserved Mycobacterium tuberculosis transcription-repair coupling factor.

Similar Datasets

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