Cleavage of the moaX-encoded fused molybdopterin synthase from Mycobacterium tuberculosis is necessary for activity

 

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dc.contributor.author Narrandes, Nicole C
dc.contributor.author Machowski, Edith E
dc.contributor.author Mizrahi, Valerie
dc.contributor.author Kana, Bavesh D
dc.date.accessioned 2015-03-04T09:46:06Z
dc.date.available 2015-03-04T09:46:06Z
dc.date.issued 2015-02-06
dc.identifier.citation Narrandes, Nicole C.; Machowski, Edith E.; Mizrahi, Valerie and Kana, Bavesh D. (2015) Cleavage of the moaX-encoded fused molybdopterin synthase from Mycobacterium tuberculosis is necessary for activity. BMC Microbiology. 15(1):22-34. en_ZA
dc.identifier.uri http://hdl.handle.net/11427/12552
dc.identifier.uri http://dx.doi.org/10.1186/s12866-015-0355-2
dc.description.abstract Background: Molybdopterin cofactor (MoCo) biosynthesis in Mycobacterium tuberculosis is associated with a multiplicity of genes encoding several enzymes in the pathway, including the molybdopterin (MPT) synthase, a hetero tetramer comprising two MoaD and two MoaE subunits. In addition to moaD1, moaD2, moaE1, moaE2, the M. tuberculosis genome also contains a moaX gene which encodes an MPT-synthase in which the MoaD and MoaE domains are located on a single polypeptide. In this study, we assessed the requirement for post-translational cleavage of MoaX for functionality of this novel, fused MPT synthase and attempted to establish a functional hierarchy for the various MPT-synthase encoding genes in M. tuberculosis. Results: Using a heterologous Mycobacterium smegmatis host and the activity of the MoCo-dependent nitrate reductase, we confirmed that moaD2 and moaE2 from M. tuberculosis together encode a functional MPT synthase. In contrast, moaD1 displayed no functionality in this system, even in the presence of the MoeBR sulphurtransferase, which contains the rhodansese-like domain, predicted to activate MoaD subunits. We demonstrated that cleavage of MoaX into its constituent MoaD and MoaE subunits was required for MPT synthase activity and confirmed that cleavage occurs between the Gly82 and Ser83 residues in MoaX. Further analysis of the Gly81-Gly82 motif confirmed that both of these residues are necessary for catalysis and that the Gly81 was required for recognition/cleavage of MoaX by an as yet unidentified protease. In addition, the MoaE component of MoaX was able to function in conjunction with M. smegmatis MoaD2 suggesting that cleavage of MoaX renders functionally interchangeable subunits. Expression of MoaX in E. coli revealed that incorrect post-translational processing is responsible for the lack of activity of MoaX in this heterologous host. Conclusions: There is a degree of functional interchangeability between the MPT synthase subunits of M. tuberculosis. In the case of MoaX, post-translational cleavage at the Gly82 residue is required for function. en_ZA
dc.language eng en_ZA
dc.publisher BioMed Central en_ZA
dc.rights Creative Commons Attribution 4.0 International (CC BY 4.0) *
dc.rights.uri http://creativecommons.org/licenses/by/4.0/ en_ZA
dc.source BMC Microbiology en_ZA
dc.source.uri http://www.biomedcentral.com/bmcmicrobiol/
dc.subject.lcsh Mycobacteria en_ZA
dc.subject.other MoCo biosynthesis en_ZA
dc.subject.other Molybdopterin synthase en_ZA
dc.subject.other MoaX en_ZA
dc.subject.other MoaD en_ZA
dc.subject.other MoaE en_ZA
dc.subject.other Nitrate reductase en_ZA
dc.title Cleavage of the moaX-encoded fused molybdopterin synthase from Mycobacterium tuberculosis is necessary for activity en_ZA
dc.type Journal Article en_ZA
dc.date.updated 2015-02-12T19:02:17Z
dc.language.rfc3066 en
dc.rights.holder Narrandes et al.; licensee BioMed Central.
uct.type.publication Research en_ZA
uct.type.resource Article en_ZA
dc.publisher.institution University of Cape Town
dc.publisher.faculty Faculty of Health Sciences en_ZA
dc.publisher.department Division of Medical Microbiology en_ZA
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uct.type.filetype Text
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