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|Title:||Identification, Characterization, and Azole-Binding Properties of Mycobacterium smegmatis CYP164A2, a Homolog of ML2088, the Sole Cytochrome P450 Gene of Mycobacterium leprae.|
|Authors:||Warrilow, Andrew G. S.|
Jackson, Colin J.
Parker, Josie E.
Marczylo, Timothy H.
Kelly, Diane E.
Lamb, David C.
Kelly, Steven L.
|Publisher:||American Society for Microbiology|
|Citation:||Antimicrobial Agents and Chemotherapy, 2009, 53 (3), pp. 1157-1164.|
|Abstract:||The genome sequence of Mycobacterium leprae revealed a single open reading frame, ML2088 (CYP164A1), encoding a putative full-length cytochrome P450 monooxygenase and 12 pseudogenes. We have identified a homolog of ML2088 in Mycobacterium smegmatis and report here the cloning, expression, purification, and azole-binding characteristics of this cytochrome P450 (CYP164A2). CYP164A2 is 1,245 bp long and encodes a protein of 414 amino acids and molecular mass of 45 kDa. CYP164A2 has 60% identity with Mycobacterium leprae CYP161A1 and 66 to 69% identity with eight other mycobacterial CYP164A1 homologs, with three identified highly conserved motifs. Recombinant CYP164A2 has the typical spectral characteristics of a cytochrome P450 monooxygenase, predominantly in the ferric low-spin state. Unusually, the spin state was readily modulated by increasing ionic strength at pH 7.5, with 50% high-spin occupancy achieved with 0.14 M NaCl. CYP164A2 bound clotrimazole, econazole, and miconazole strongly (Kd, 1.2 to 2.5 µM); however, strong binding with itraconazole, ketoconazole, and voriconazole was only observed in the presence of 0.5 M NaCl. Fluconazole did not bind to CYP164A2 at pH 7.5 and no discernible type II binding spectrum was observed.|
|Rights:||This is the author's final draft of the paper published as Antimicrobial Agents and Chemotherapy, 2009, 53 (3), pp. 1157-1164. The final version is available from http://aac.asm.org/cgi/content/abstract/53/3/1157. Doi: 10.1128/AAC.01237-08|
|Appears in Collections:||Published Articles, Dept. of Cancer Studies and Molecular Medicine|
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