title: How does the reductase help to regulate the catalytic cycle of cytochrome P450 3A4 using the conserved water channel?
abstract: Water molecules play a major role in the P450 catalytic cycle. Here, we locate the preferred water pathways and their gating mechanisms for the human cytochrome P450 3A4 (CYP3A4) and elucidate the role of the cytochrome P450 reductase (CPR) in turning on and activating these water channels. We perform explicit solvent molecular dynamic simulations of CYP3A4, unbound and bound to two substrates, and with and without the flavin mononucleotide (FMN)-binding domain of CPR. We observe in/out passage of water molecules via a water-specific and conserved channel (aqueduct) located between the active site and the heme proximal side. We find that the aqueduct gating mechanism is mediated by R375, the conserved arginine that salt bridges with the heme 7-propionate. When R375 rotates, it opens the aqueduct and establishes a connection between a cluster of active site water molecules network and the bulk solvent. The aqueduct region overlaps with the CPR binding-site to CYP3A4. Indeed, we find that when the FMN domain of CPR binds to CYP3A4, the aqueduct fully opens up, thereby allowing a flow of water molecules. The aqueduct's opening can permit proton transfer, shuttling the protons to the active site through ordered water molecules. In addition, the expulsion of water molecules via the aqueduct contributes to substrate binding. As such, the CPR binding has a function: it triggers the aqueduct's opening and thereby enables a proton shuttle pathway, which is needed for the dioxygen activation. This mechanism could be a general paradigm in P450s.
authors.affiliation: Department of Human Genetics, Sackler Institute of Molecular Medicine, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Dan
authors.initials: D
authors.lastname: Fishelovitch
authors.forename: Sason
authors.initials: S
authors.lastname: Shaik
authors.forename: Haim J
authors.initials: HJ
authors.lastname: Wolfson
authors.forename: Ruth
authors.initials: R
authors.lastname: Nussinov
mesh_terms: D055162:Biocatalysis; D020134:Catalytic Domain; D051544:Cytochrome P-450 CYP3A / Q000737:chemistry*; D006801:Humans; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D014867:Water / Q000737:chemistry*
chemical_list: D014867:Water; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human
references.citation: J Mol Biol. 1995 Apr 7;247(4):536-40
references.pmid: 7723011
references.citation: J Am Chem Soc. 2008 Jan 16;130(2):432-3
references.pmid: 18088124
references.citation: Biochemistry. 2001 Nov 13;40(45):13456-65
references.pmid: 11695892
references.citation: Drug Metab Dispos. 1999 Sep;27(9):999-1004
references.pmid: 10460798
references.citation: J Biol Chem. 2006 Mar 17;281(11):7614-22
references.pmid: 16352597
references.citation: J Comput Chem. 2005 Dec;26(16):1781-802
references.pmid: 16222654
references.citation: J Biol Chem. 1998 Jul 3;273(27):17036-49
references.pmid: 9642268
references.citation: J Phys Chem B. 2007 May 17;111(19):5445-57
references.pmid: 17441761
references.citation: Proc Natl Acad Sci U S A. 2005 Jul 26;102(30):10478-81
references.pmid: 16014708
references.citation: J Am Chem Soc. 2007 Feb 14;129(6):1602-11
references.pmid: 17284003
references.citation: J Biol Chem. 2007 Oct 12;282(41):29766-76
references.pmid: 17693640
references.citation: J Mol Biol. 1994 Mar 4;236(4):1169-85
references.pmid: 8120894
references.citation: Biochemistry. 1993 Nov 2;32(43):11530-8
references.pmid: 8218220
references.citation: Chem Rev. 1996 Nov 7;96(7):2841-2888
references.pmid: 11848843
references.citation: Drug Metab Rev. 2000 Aug-Nov;32(3-4):339-61
references.pmid: 11139133
references.citation: Proc Natl Acad Sci U S A. 2003 Nov 11;100(23):13121-2
references.pmid: 14597705
references.citation: J Phys Chem B. 2005 Jan 27;109(3):1268-80
references.pmid: 16851091
references.citation: Biochemistry. 1998 Jun 30;37(26):9211-9
references.pmid: 9649301
references.citation: Arch Biochem Biophys. 2000 Nov 1;383(1):70-8
references.pmid: 11097178
references.citation: J Comput Chem. 2004 Aug;25(11):1400-15
references.pmid: 15185334
references.citation: Biochim Biophys Acta. 2007 Mar;1770(3):390-401
references.pmid: 16920266
references.citation: J Biol Chem. 2005 Dec 23;280(51):42188-97
references.pmid: 16239228
references.citation: Nucleic Acids Res. 2008 Jul 1;36(Web Server issue):W210-5
references.pmid: 18448468
references.citation: Science. 2004 Jul 30;305(5684):683-6
references.pmid: 15256616
references.citation: Science. 2000 Mar 3;287(5458):1615-22
references.pmid: 10698731
references.citation: J Phys Chem B. 1998 Apr 30;102(18):3586-616
references.pmid: 24889800
references.citation: J Am Chem Soc. 2009 Apr 8;131(13):4755-63
references.pmid: 19281168
references.citation: Biochemistry. 2001 Mar 6;40(9):2669-77
references.pmid: 11258878
references.citation: Proc Natl Acad Sci U S A. 1997 Mar 18;94(6):2133-8
references.pmid: 9122160
references.citation: Biophys J. 2006 Nov 15;91(10):3630-9
references.pmid: 16935961
references.citation: Proteins. 2006 Jan 1;62(1):209-17
references.pmid: 16294339
references.citation: Nat Struct Biol. 1996 May;3(5):414-7
references.pmid: 8612070
references.citation: J Comput Chem. 2005 Oct;26(13):1383-99
references.pmid: 16028234
references.citation: Chem Rev. 2005 Jun;105(6):2279-328
references.pmid: 15941215
references.citation: J Am Chem Soc. 2001 Feb 21;123(7):1403-15
references.pmid: 11456714
references.citation: Proc Natl Acad Sci U S A. 2006 Sep 12;103(37):13682-7
references.pmid: 16954191
references.citation: Biochemistry. 1997 Apr 15;36(15):4461-70
references.pmid: 9109653
references.citation: J Biol Chem. 2004 Sep 10;279(37):38091-4
references.pmid: 15258162
references.citation: Eur J Biochem. 1977 Nov 1;80(2):319-24
references.pmid: 923582
references.citation: J Am Chem Soc. 2009 Feb 4;131(4):1398-400
references.pmid: 19133773
references.citation: Biochemistry. 2007 Dec 25;46(51):14838-44
references.pmid: 18044969
references.citation: Proc Natl Acad Sci U S A. 1999 Mar 2;96(5):1863-8
references.pmid: 10051560
references.citation: J Biol Chem. 2005 Sep 9;280(36):31659-63
references.pmid: 15994329
references.citation: Biophys J. 2002 Oct;83(4):1731-48
references.pmid: 12324397
references.citation: Bioinformatics. 2004 May 22;20(8):1322-4
references.pmid: 14871869
references.citation: Nucleic Acids Res. 2005 Jul 1;33(Web Server issue):W363-7
references.pmid: 15980490
references.citation: Drug Discov Today. 2004 Jun 15;9(12):530-7
references.pmid: 15183161