Homogeneous recombinant Mycobacterium tuberculosis shikimate dehydrogenase production: an essential step towards target-based drug design.
Mycobacterium tuberculosis shikimate dehydrogenase (MtbSD) catalyzes the forth reaction in the shikimate pathway. Here we describe production of K69A, K69H, K69I, K69Q, D105A, and D105N mutant proteins. Screening of several conditions was performed to optimize MtbSD production yield, and an improved purification protocol to obtain homogeneous MtbSD is presented. The rational design of new antitubercular drugs hinges on the availability of M. tuberculosis proteins. Our results show that optimization of expression, disruption, and purification protocols resulted in a higher yield of functional MtbSD enzyme.
Rodrigues
Valnês da Silva
Vda S
Centro de Pesquisas em Biologia Molecular e Funcional, Instituto Nacional de Ciência e Tecnologia em Tuberculose, Pontifícia Universidade Católica do Rio Grande do Sul (PUCRS), Porto Alegre, RS 90619-900, Brazil.
Basso
Luiz Augusto
LA
Santos
Diógenes Santiago
DS
D000429:Alcohol Oxidoreductases / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D000900:Anti-Bacterial Agents / Q000378:metabolism / Q000494:pharmacology; D015195:Drug Design; D004591:Electrophoresis, Polyacrylamide Gel; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000187:drug effects / Q000201:enzymology*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D016415:Sequence Alignment
D000900:Anti-Bacterial Agents; D011994:Recombinant Proteins; D000429:Alcohol Oxidoreductases; C022641:Shikimate dehydrogenase