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Biocatalytic preparation and characterization of alternative substrate of MshB, a mycothiol pathway enzyme
Thesis (MSc)--Stellenbosch University, 2012.
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| Format: | Thesis |
| Language: | en_ZA |
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Stellenbosch : Stellenbosch University
2012
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| _version_ | 1867614098096652288 |
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| access_status_str | Open Access |
| author | Muneri, Ndivhuwo Olga |
| author2 | Strauss, Erick |
| author_browse | Muneri, Ndivhuwo Olga Strauss, Erick |
| author_facet | Strauss, Erick Muneri, Ndivhuwo Olga |
| author_sort | Muneri, Ndivhuwo Olga |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Thesis (MSc)--Stellenbosch University, 2012. |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/71972 |
| institution | Stellenbosch University (South Africa) |
| language | en_ZA |
| last_indexed | 2026-06-10T12:46:38.139Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2012 |
| publishDateRange | 2012 |
| publishDateSort | 2012 |
| publisher | Stellenbosch : Stellenbosch University |
| publisherStr | Stellenbosch : Stellenbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/71972 Biocatalytic preparation and characterization of alternative substrate of MshB, a mycothiol pathway enzyme Muneri, Ndivhuwo Olga Strauss, Erick Stellenbosch University. Faculty of Science. Dept. of Biochemistry. Mycothiol pathway enzyme MshB -- Biocatalytic preparation MshB -- characterisation UCTD Thesis (MSc)--Stellenbosch University, 2012. ENGLISH ABSTRACT: Mycobacterium tuberculosis (M. tuberculosis), the causative agent of tuberculosis, utilizes mycothiol (MSH) as the major low molecular weight thiol to protect itself against oxidative stress and thereby to ensure its growth and survival. MSH is a pseudo-disaccharide molecule that contains an α(1→1) glycosidic bond, and is biosynthesised in five enzymatic steps involving the enzymes MshA, MshA2, MshB, MshC and MshD. Owing to the essentiality of MSH to M. tuberculosis, various studies have focused on the MSH biosynthetic and other MSH-dependent enzymes viewed as potential drug targets for the development of antituberculosis agents. In the course of this study two practical challenges affecting the development of inhibitors of one the MSH biosynthesis pathway enzyme, MshB, were addressed. These challenges entail the lack of a high-throughput continuous assay to determine MshB activity, and the poor availability of the natural and alternative MshB substrates. In this study an alternate MshB substrate was characterized and shown to undergo a rearrangement reaction upon deactylation, which allowed the development of a new continuous assay for MshB activity that uses DNTB (Ellman’s reagent). In addition, three new α-thioglycoligases were created from the α-Nacetylglucosaminidase of Clostridium perfringens. These enzymes showed potential as biocatalysts that can be used for the enzymatic synthesis of thioglycoside-based alternative substrates of MshB. AFRIKAANSE OPSOMMING: Mycobacterium tuberculosis, die organisme wat tuberkulose veroorsaak, maak gebruik van mikotiol (MSH) om homself te beskerm teen oksidatiewe stres en sodoende sy groei en oorlewing te verseker. MSH is ʼn pseudo-disakkaried molekule met ʼn α(1→1) glikosidiese binding, en word in vyf ensimatiese stappe gebiosintetiseer deur die ensieme MshA, MshA2, MshB, MshC en MshD. Weens die noodsaaklikheid van MSH vir M. tuberculosis het verskeie vorige studies gefokus op die MSH biosintetiese en ander MSH-afhanklike ensieme as potensiële teikens vir die ontwikkeling van antituberkulose middels. In die loop van hierdie studie is twee praktiese uitdagings wat die ontwikkeling van inhibitors van een van die MSH biosintetiese ensieme, MshB, bemoeilik, aangespreek. Hierdie uitdagings behels die gebrek aan 'n geskikte hoë-deurvloei kontinue essaï vir MshB aktiwiteit, en die lae beskikbaarheid van die natuurlike en alternatiewe MshB substrate. In hierdie studie is alternatiewe MshB substrate gekarakteriseer en is daar gewys dat dit ʼn herrangskikkingsreaksie ondergaan na deasetilering, wat die ontwikkeling van ʼn nuwe kontinue essaï vir MshB aktiwiteit wat gebruik maak van DTNB (Ellman se reagens), moontlik gemaak het. Verder is drie nuwe α-tioglikoligases ontwikkel van die ensiem α-N-asetielglukosaminidase van Clostridium perfringens. Hierdie ensieme toon potensiaal as biokataliste wat gebruik kan word in die ensiematiese sintese van tioglikosied-gebaseerde alternatiewe substrate van MshB. 2012-11-28T15:08:09Z 2012-12-12T08:19:37Z 2012-11-28T15:08:09Z 2012-12-12T08:19:37Z 2012-12 Thesis http://hdl.handle.net/10019.1/71972 en_ZA Stellenbosch University 98 pages : illustrations application/pdf application/pdf Stellenbosch : Stellenbosch University |
| spellingShingle | Mycothiol pathway enzyme MshB -- Biocatalytic preparation MshB -- characterisation UCTD Muneri, Ndivhuwo Olga Biocatalytic preparation and characterization of alternative substrate of MshB, a mycothiol pathway enzyme |
| title | Biocatalytic preparation and characterization of alternative substrate of MshB, a mycothiol pathway enzyme |
| title_full | Biocatalytic preparation and characterization of alternative substrate of MshB, a mycothiol pathway enzyme |
| title_fullStr | Biocatalytic preparation and characterization of alternative substrate of MshB, a mycothiol pathway enzyme |
| title_full_unstemmed | Biocatalytic preparation and characterization of alternative substrate of MshB, a mycothiol pathway enzyme |
| title_short | Biocatalytic preparation and characterization of alternative substrate of MshB, a mycothiol pathway enzyme |
| title_sort | biocatalytic preparation and characterization of alternative substrate of mshb a mycothiol pathway enzyme |
| topic | Mycothiol pathway enzyme MshB -- Biocatalytic preparation MshB -- characterisation UCTD |
| url | http://hdl.handle.net/10019.1/71972 |
| work_keys_str_mv | AT munerindivhuwoolga biocatalyticpreparationandcharacterizationofalternativesubstrateofmshbamycothiolpathwayenzyme |