The Role of Protein-Protein and Protein-Membrane Interactions on P450 Function

Abstract

This symposium summary, sponsored by the ASPET, was held at Experimental Biology 2015 on March 29, 2015, in Boston, Massachusetts. The symposium focused on: 1) the interactions of cytochrome P450s (P450s) with their redox partners; and 2) the role of the lipid membrane in their orientation and stabilization. Two presentations discussed the interactions of P450s with NADPH-P450 reductase (CPR) and cytochrome b 5 . First, solution nuclear magnetic resonance was used to compare the protein interactions that facilitated either the hydroxylase or lyase activities of CYP17A1. The lyase interaction was stimulated by the presence of b 5 and 17a-hydroxypregnenolone, whereas the hydroxylase reaction was predominant in the absence of b 5 . The role of b 5 was also shown in vivo by selective hepatic knockout of b 5 from mice expressing CYP3A4 and CYP2D6; the lack of b 5 caused a decrease in the clearance of several substrates. The role of the membrane on P450 orientation was examined using computational methods, showing that the proximal region of the P450 molecule faced the aqueous phase. The distal region, containing the substrate-access channel, was associated with the membrane. The interaction of NADPH-P450 reductase (CPR) with the membrane was also described, showing the ability of CPR to "helicopter" above the membrane. Finally, the endoplasmic reticulum (ER) was shown to be heterogeneous, having ordered membrane regions containing cholesterol and more disordered regions. Interestingly, two closely related P450s, CYP1A1 and CYP1A2, resided in different regions of the ER. The structural characteristics of their localization were examined. These studies emphasize the importance of P450 protein organization to their function.

Cytochrome b5 Is an Obligate Electron Shuttle Protein for Syringyl Lignin Biosynthesis in Arabidopsis

“…whether the copurified proteins were in direct physical interactions or in spatial proximity with the baits, our study revealed several electron transfer proteins that coeluted with monolignol P450 enzymes (Table 1). This discovery coincides with the previous demonstration that mammalian and human microsomal P450s physically interact with their redox partners, CPR and CB5, via electrostatic interactions at the basic, positively charged proximal surfaces of P450s (Schenkman and Jansson, 1999;Im and Waskell, 2011;Kandel and Lampe, 2014;Scott et al, 2016). The interaction of P450 with its electron carrier was proposed to form a functional 1:1 complex (Miwa and Lu, 1984).…”

“…Cytochrome P450 enzymes physically and stochastically associate with their redox partner CPR or CB5 via electrostatic interactions to form functional 1:1 complexes (Vergéres and Waskell, 1995;Backes and Kelley, 2003;Im and Waskell, 2011;Scott et al, 2016). Such redox partner-enzyme complex formation is essential for the roles of P450 enzymes in xenobiotic detoxification and drug metabolism in mammals (Kandel and Lampe, 2014).…”

The Hinge Segment of Human NADPH-Cytochrome P450 Reductase in Conformational Switching: The Critical Role of Ionic Strength

“…The catalytic cycle of P450 enzymes depends on redox partners for electron delivery. The NADPH-cytochrome P450 reductase (CPR) is strictly required for the activity of microsomal P450s, while cytochrome b 5 can potentially be a donor molecule, albeit only for the second electron transfer step ( Scott et al, 2016 ).…”

Interaction of enzymes with lignocellulosic materials: causes, mechanism and influencing factors

“…Three types of interactions are involved in adsorption of enzyme onto lignin: (i) hydrophobic interactions (Ying et al 2018;Tokunaga et al 2019); (ii) electrostatic interactions (Nakagame et al 2011;Scott et al 2016), and (iii) hydrogen-bonding (Rahikainen et al 2013;Liu et al 2016). The CBDs of T. reesei cellulases, Cel7A and Cel5A were adsorbed onto lignin in significantly high amount (Yarbrough, et al 2015;Liu et al 2016) due to hydrophobic interaction.…”