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Competitive inhibitors of Mycobacterium tuberculosis ribose-5-phosphate isomerase B reveal new information about the reaction mechanism.
Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology.
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2005 In: J Biol Chem, ISSN 0021-9258, Vol. 280, no 8, 6416-6422 p.Article in journal (Refereed) Published
Place, publisher, year, edition, pages
2005. Vol. 280, no 8, 6416-6422 p.
Identifiers
URN: urn:nbn:se:uu:diva-96133OAI: oai:DiVA.org:uu-96133DiVA: diva2:170607
Available from: 2007-09-07 Created: 2007-09-07Bibliographically approved
In thesis
1. Structural and Functional Studies of Ribose-5-phosphate isomerase B
Open this publication in new window or tab >>Structural and Functional Studies of Ribose-5-phosphate isomerase B
2007 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Ribose 5-phosphate isomerase (Rpi) is one of the major enzymes of the pentose phosphate pathway, where it catalyses the inter-conversion of ribose 5-phosphate (R5P) and ribulose 5-phosphate. Two forms of this isomerase with no significant amino acid sequence similarity exist, RpiA and RpiB. This thesis describes RpiB from the organisms Mycobacterium tuberculosis (Mt) and Escherichia coli (Ec) from a structural and functional point of view.

Since the E. coli genome encodes both an RpiA and an RpiB, which generally is not expressed, it has been proposed that EcRpiB has a different role as an allose-6-phosphate isomerase. Activity measurements presented here show that EcRpiB does have this second activity.

In the M. tuberculosis genome there is only a gene for RpiB. The crystal structure of MtRpiB was solved in complex with several different inhibitors designed to mimic the reaction intermediate as well as with the substrate, R5P. The organisation of the active site in these structures could be used to derive the reaction mechanism for MtRpiB and for other RpiBs in general. Activity measurements of MtRpiB showed that it can catalyse the R5P isomerisation, but not the allose 6-phosphate reaction. Differences observed in the active site between EcRpiB and MtRpiB explain these kinetic results.

Activity measurements and a structure of an EcRpiB mutant, where histidine99 was changed to asparagine, implies that RpiB catalyses the first step of the reaction in which the sugar ring must be opened, and gives a possible explanation for how this could occur.

Inhibition studies have uncovered a compound that selectively inhibits MtRpiB over RpiA from spinach, which is homologous to the human RpiA. Differences in the inhibition patterns and active site residues of these two species’ Rpi may provide information for future virtual screening approaches, with the aim of discovering new anti-tuberculosis agents.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2007. iv, 73 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 332
Keyword
Molecular biology, Mycobacterium tuberculosis, reaction mechanism, cis-enediolate high energy intermediate, allose-6-phosphate isomerase, rare sugar, pentose phosphate pathway, Rv2465c, X-ray crystallography, Molekylärbiologi
Identifiers
urn:nbn:se:uu:diva-8182 (URN)978-91-554-6952-8 (ISBN)
Public defence
2007-09-28, B42, BMC, Husargatan 3, Uppsala, 13:00
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Available from: 2007-09-07 Created: 2007-09-07Bibliographically approved

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