Journal article

Publication year: 2006

Pages: 32967-32977

Journal: Journal of Biological Chemistry

Volume number: 281

Issue number: 44

ISSN: 0021-9258

eISSN: 1083-351X

Open access status: Hybrid

DOI Link: https://doi.org/10.1074/jbc.M601141200

Publisher: Elsevier

Abstract: 
High-M-r thioredoxin reductase from the malaria parasite Plasmodium falciparum (PfTrxR) contains three redox active centers (FAD, Cys-88/Cys-93, and Cys-535/Cys-540) that are in redox communication. The catalytic mechanism of PfTrxR, which involves dithiol-disulfide interchanges requiring acid-base catalysis, was studied by steady-state kinetics, spectral analyses of anaerobic static titrations, and rapid kinetics analysis of wild-type enzyme and variants involving the His-509-Glu-514 dyad as the presumed acid-base catalyst. The dyad is conserved in all members of the enzyme family. Substitution of His-509 with glutamine and Glu-514 with alanine led to TrxR with only 0.5 and 7% of wild type activity, respectively, thus demonstrating the crucial roles of these residues for enzymatic activity. The H509Q variant had rate constants in both the reductive and oxidative half-reactions that were dramatically less than those of wild-type enzyme, and no thiolate-flavin charge-transfer complex was observed. Glu-514 was shown to be involved in dithiol-disulfide interchange between the Cys-88/Cys-93 and Cys-535/Cys-540 pairs. In addition, Glu-514 appears to greatly enhance the role of His-509 in acid-base catalysis. It can be concluded that the His-509-Glu-514 dyad, in analogy to those in related oxidoreductases, acts as the acid-base catalyst in PfTrxR.

Harvard Citation style: McMillan, P., Arscott, L., Ballou, D., Becker, K., Williams, C. and Müller, S. (2006) Identification of acid-base catalytic residues of high-M-r thioredoxin reductase from Plasmodium falciparum, Journal of Biological Chemistry, 281(44), pp. 32967-32977. https://doi.org/10.1074/jbc.M601141200

APA Citation style: McMillan, P., Arscott, L., Ballou, D., Becker, K., Williams, C., & Müller, S. (2006). Identification of acid-base catalytic residues of high-M-r thioredoxin reductase from Plasmodium falciparum. Journal of Biological Chemistry. 281(44), 32967-32977. https://doi.org/10.1074/jbc.M601141200