Thiamine-phosphate kinase (or ATP:thiamine-phosphate phosphotransferase, ThiL) and
homoserine (ThrB) kinases are essential to metabolism in Mycobacterium tuberculosis
(Mtb). ThiL and ThrB respectively phosphorylate thiamine monophosphate (TMP) to
thiamine diphosphate (TDP), the active form of vitamin B1, and L-homoserine to Ophosphohomoserine,
critical to aspartate biosynthesis.
In this study, ThiL and ThrB from Mtb were characterised structurally and functionally by
producing the proteins recombinantly in E. coli. Proteins were purified by affinity, anion
exchange and size exclusion chromatographies and purity checked by SDS-PAGE. ThiL and
ThrB enzyme activities were confirmed and reaction products verified by high pressure
liquid chromatography (HPLC). The crystal structure of ThiL was solved by molecular
replacement using X-ray diffraction data.
Functionally active ThiL, 36 kDa, produced hexagonal crystals belonging to space group
P6122 with one monomer per asymmetric unit. Structurally it is related to ThiL from other
organisms with minor structural deviations.
Enzymatically active ThrB, 33 kDa, was crystallised. However, crystals failed to diffract Xrays
to a suitable resolution. ThiL and ThrB could act as possible anti-TB drug targets
against Mtb. / Dissertation (MSc)--University of Pretoria, 2017. / Biochemistry / MSc / Unrestricted
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:up/oai:repository.up.ac.za:2263/63284 |
| Date | January 2017 |
| Creators | Ntui, Clifford Manyo |
| Contributors | Schubert, Wolf-Dieter, u14334012@tuks.co.za, Motshwene, Precious, G. |
| Publisher | University of Pretoria |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Dissertation |
| Rights | © 2017 University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. |
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