TY - JOUR
T1 - Mechanistic scrutiny identifies a kinetic role for cytochrome b5 regulation of human cytochrome P450c17 (CYP17A1, P450 17A1)
AU - Simonov, Alexandr
AU - Holien, Jessica K
AU - Yeung, Chun In
AU - Nguyen, Ann D
AU - Corbin, Cynthia Jo
AU - Zheng, Jie
AU - Kuznetsov, Vladimir L
AU - Auchus, Richard Joshph
AU - Conley, Alan James
AU - Bond, Alan Maxwell
AU - Parker, Michael William
AU - Rodgers, Raymond J
AU - Martin, Lisandra Lorraine
PY - 2015
Y1 - 2015
N2 - Cytochrome P450c17 (P450 17A1, CYP17A1) is a critical enzyme in the synthesis of androgens and is now a target enzyme for the treatment of prostate cancer. Cytochrome P450c17 can exhibit either one or two physiological enzymatic activities differentially regulated by cytochrome b5. How this is achieved remains unknown. Here, comprehensive in silico, in vivo and in vitro analyses were undertaken. Fluorescence Resonance Energy Transfer analysis showed close interactions within living cells between cytochrome P450c17 and cytochrome b5. In silico modeling identified the sites of interaction and confirmed that E48 and E49 residues in cytochrome b5 are essential for activity. Quartz crystal microbalance studies identified specific protein-protein interactions in a lipid membrane. Voltammetric analysis revealed that the wild type cytochrome b5, but not a mutated, E48G/E49G cyt b5, altered the kinetics of electron transfer between the electrode and the P450c17. We conclude that cytochrome b5 can influence the electronic conductivity of cytochrome P450c17 via allosteric, protein-protein interactions
AB - Cytochrome P450c17 (P450 17A1, CYP17A1) is a critical enzyme in the synthesis of androgens and is now a target enzyme for the treatment of prostate cancer. Cytochrome P450c17 can exhibit either one or two physiological enzymatic activities differentially regulated by cytochrome b5. How this is achieved remains unknown. Here, comprehensive in silico, in vivo and in vitro analyses were undertaken. Fluorescence Resonance Energy Transfer analysis showed close interactions within living cells between cytochrome P450c17 and cytochrome b5. In silico modeling identified the sites of interaction and confirmed that E48 and E49 residues in cytochrome b5 are essential for activity. Quartz crystal microbalance studies identified specific protein-protein interactions in a lipid membrane. Voltammetric analysis revealed that the wild type cytochrome b5, but not a mutated, E48G/E49G cyt b5, altered the kinetics of electron transfer between the electrode and the P450c17. We conclude that cytochrome b5 can influence the electronic conductivity of cytochrome P450c17 via allosteric, protein-protein interactions
UR - http://journals.plos.org/plosone/article/asset?id=10.1371%2Fjournal.pone.0141252.PDF
U2 - 10.1371/journal.pone.0141252
DO - 10.1371/journal.pone.0141252
M3 - Article
VL - 10
SP - 1
EP - 19
JO - PLoS ONE
JF - PLoS ONE
SN - 1932-6203
IS - 11
M1 - e0141252
ER -