Characterization of active site structure in CYP121: A cytochrome P450 essential for viability of mycobacterium tuberculosis H37Rv

McLean, Kirsty J., Carroll, Paul, Lewis, D. Geraint, Dunford, Adrian J., Seward, Harriet E., Neeli, Rajasekhar, Cheesman, Myles R., Marsollier, Laurent, Douglas, Philip, Smith, W. Ewen, Rosenkrands, Ida, Cole, Stewart T., Leys, David, Parish, Tanya and Munro, Andrew W. (2008) Characterization of active site structure in CYP121: A cytochrome P450 essential for viability of mycobacterium tuberculosis H37Rv. Journal of Biological Chemistry, 283 (48). pp. 33406-33416. ISSN 0021-9258

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Abstract

Mycobacterium tuberculosis (Mtb) cytochrome P450 gene CYP121 is shown to be essential for viability of the bacterium in vitro by gene knock-out with complementation. Production of CYP121 protein in Mtb cells is demonstrated. Minimum inhibitory concentration values for azole drugs against Mtb H37Rv were determined, the rank order of which correlated well with K-d values for their binding to CYP121. Solution-state spectroscopic, kinetic, and thermodynamic studies and crystal structure determination for a series of CYP121 active site mutants provide further insights into structure and biophysical features of the enzyme. Pro(346) was shown to control heme cofactor conformation, whereas Arg(386) is a critical determinant of heme potential, with an unprecedented 280-mV increase in heme iron redox potential in a R386L mutant. A homologous Mtb redox partner system was reconstituted and transported electrons faster to CYP121 R386L than to wild type CYP121. Hemepotential was not perturbed in a F338H mutant, suggesting that a proposed P450 superfamily-wide role for the phylogenetically conserved phenylalanine in heme thermodynamic regulation is unlikely. Collectively, data point to an important cellular role for CYP121 and highlight its potential as a novel Mtb drug target.

Item Type:	Article
Uncontrolled Keywords:	sdg 3 - good health and well-being ,/dk/atira/pure/sustainabledevelopmentgoals/good_health_and_well_being
Faculty \ School:	Faculty of Science > School of Chemistry (former - to 2024)
UEA Research Groups:	Faculty of Science > Research Groups > Biophysical Chemistry (former - to 2017) Faculty of Science > Research Groups > Chemistry of Life Processes Faculty of Science > Research Centres > Centre for Molecular and Structural Biochemistry Faculty of Science > Research Groups > Chemistry of Light and Energy
Depositing User:	Rachel Smith
Date Deposited:	10 May 2011 10:58
Last Modified:	24 Sep 2024 09:33
URI:	https://ueaeprints.uea.ac.uk/id/eprint/30081
DOI:	10.1074/jbc.M802115200

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