Effect of homomeric P450-P450 complexes on P450 function.
Previous studies have shown that the presence of one P450 enzyme can affect the function of another. The goal of the present study was to determine if P450 enzymes are capable of forming homomeric complexes that affect P450 function. To address this problem, the catalytic activities of several P450s were examined in reconstituted systems containing NADPH-POR (cytochrome P450 reductase) and a single P450. CYP2B4 (cytochrome P450 2B4)-, CYP2E1 (cytochrome P450 2E1)- and CYP1A2 (cytochrome P450 1A2)-mediated activities were measured as a function of POR concentration using reconstituted systems containing different concentrations of P450. Although CYP2B4-dependent activities could be explained by a simple Michaelis-Menten interaction between POR and CYP2B4, both CYP2E1 and CYP1A2 activities generally produced a sigmoidal response as a function of [POR]. Interestingly, the non-Michaelis behaviour of CYP1A2 could be converted into a simple mass-action response by increasing the ionic strength of the buffer. Next, physical interactions between CYP1A2 enzymes were demonstrated in reconstituted systems by chemical cross-linking and in cellular systems by BRET (bioluminescence resonance energy transfer). Cross-linking data were consistent with the kinetic responses in that both were similarly modulated by increasing the ionic strength of the surrounding solution. Taken together, these results show that CYP1A2 forms CYP1A2-CYP1A2 complexes that exhibit altered catalytic activity.
Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Sciences Center, 533 Bolivar Street, New Orleans, LA 70112, USA.
James R
JR
Reed
J Patrick
JP
Connick
Dongmei
D
Cheng
George F
GF
Cawley
Wayne L
WL
Backes
D000818:Animals; D060807:Bioluminescence Resonance Energy Transfer Techniques; D002384:Catalysis; D019388:Cytochrome P-450 CYP1A2 / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D054730:Protein Interaction Domains and Motifs; D011817:Rabbits
D003577:Cytochrome P-450 Enzyme System; C504573:CYP1A2 protein, human; D019388:Cytochrome P-450 CYP1A2