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Structural, biochemical and in vivo investigations of the threonine synthase from Mycobacterium tuberculosis
Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Strukturell molekylärbiologi.
Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Strukturell molekylärbiologi.
Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för cell- och molekylärbiologi, Strukturell molekylärbiologi.
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2008 (Engelska)Ingår i: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 381, nr 3, s. 622-633Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Threonine biosynthesis is a general feature of prokaryotes, eukaryotic microorganisms, and higher plants. Since mammals lack the appropriate synthetic machinery, instead obtaining the amino acid through their diet, the pathway is a potential focus for the development of novel antibiotics, antifungal agents, and herbicides. Threonine synthase (TS), a pyridoxal-5-phosphate-dependent enzyme, catalyzes the final step in the pathway, in which L-homoserine phosphate and water are converted into threonine and inorganic phosphate. In the present publication, we report structural and functional studies of Mycobacterium tuberculosis TS, the product of the rv1295 (thrC) gene. The structure gives new insights into the catalytic mechanism of TSs in general, specifically by suggesting the direct involvement of the phosphate moiety of the cofactor, rather than the inorganic phosphate product, in transferring a proton from C4' to C-gamma in the formation of the alpha beta-unsaturated aldimine. It further provides a basis for understanding why this enzyme has a higher pH optimum than has been reported elsewhere for TSs and gives rise to the prediction that the equivalent enzyme from Thermus thermophilus will exhibit similar behavior. A deletion of the relevant gene generated a strain of M. tuberculosis that requires threonine for growth, such auxotrophic strains are frequently attenuated in vivo, indicating that TS is a potential drug target in this organism.

Ort, förlag, år, upplaga, sidor
2008. Vol. 381, nr 3, s. 622-633
Nyckelord [en]
threonine biosynthesis, tuberculosis, enzyme mechanism, X-ray structure, drug target
Nationell ämneskategori
Biologiska vetenskaper
Identifikatorer
URN: urn:nbn:se:uu:diva-96920DOI: 10.1016/j.jmb.2008.05.086ISI: 000258736700011PubMedID: 18621388OAI: oai:DiVA.org:uu-96920DiVA, id: diva2:171656
Tillgänglig från: 2008-03-20 Skapad: 2008-03-20 Senast uppdaterad: 2017-12-14Bibliografiskt granskad
Ingår i avhandling
1. Targeting Mycobacterium tuberculosis Proteins: Structure and Function Studies of Five Essential Proteins
Öppna denna publikation i ny flik eller fönster >>Targeting Mycobacterium tuberculosis Proteins: Structure and Function Studies of Five Essential Proteins
2008 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Abstract [en]

This thesis describes the target selection, cloning, expression, purification, crystallization, structure and biochemical characterization of five essential Mycobacterium tuberculosis (Mtb) proteins. The search for drugs against the causal agent of tuberculosis is urgently needed and the targeting of essential genes is necessary to fulfill this goal.

The crystal structures of carbonic anhydrases (CA) Rv1284 and Rv3588c have been determined to 2.0 and 1.7 Å resolution, respectively. Rv3588c, in contrast to Rv1284, is an active β-CA that shows two different active site conformations and pH-dependent oligomerization states.

Rv1295 is an active threonine synthase with an unusually high pH optimum; the structure has been solved to 2.5 Å resolution, based on which a modification to the reaction mechanism published previously is proposed.

Mtb has a thick and impermeable cell envelope that constitutes an efficient barrier against drugs. One of the essential components of the envelope is mycolic acid (MA). The inhibition of enzymes participating in its synthesis would be lethal for Mtb. Rv0636, a formerly unknown-function protein has β-hydroxyacyl-ACP dehydrase activity which is essential for MA synthesis. Co-expression with partners notably improves its solubility.

Around 55% of Mtb proteins have unknown function. Rv3778c is one of them and its three-dimensional structure has been determined to 1.8 Å resolution. Studies aimed at the elucidation of its biochemical function are shown. A pathway not yet reported in Mtb is also suggested.

Ort, förlag, år, upplaga, sidor
Uppsala: Acta Universitatis Upsaliensis, 2008. s. 61
Serie
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 411
Nyckelord
Cell and molecular biology, carbonic anhydrase, threonine synthase, FAS II, Rv0636, Rv1284, Rv1295, Rv3588c, Rv3778c, X-ray crystallography, Cell- och molekylärbiologi, Mycobacterium tuberculosis
Identifikatorer
urn:nbn:se:uu:diva-8580 (URN)978-91-554-7134-7 (ISBN)
Disputation
2008-04-11, B42, BMC, Husargatan 3, Uppsala, 13:00 (Engelska)
Opponent
Handledare
Tillgänglig från: 2008-03-20 Skapad: 2008-03-20 Senast uppdaterad: 2009-06-01Bibliografiskt granskad

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