title: Homology modeling of the three membrane proteins of the dhurrin metabolon: catalytic sites, membrane surface association and protein-protein interactions.
abstract: Formation of metabolons (macromolecular enzyme complexes) facilitates the channelling of substrates in biosynthetic pathways. Metabolon formation is a dynamic process in which transient structures mediated by weak protein-protein interactions are formed. In Sorghum, the cyanogenic glucoside dhurrin is derived from l-tyrosine in a pathway involving the two cytochromes P450 (CYPs) CYP79A1 and CYP71E1, a glucosyltransferase (UGT85B1), and the redox partner NADPH-dependent cytochrome P450 reductase (CPR). Experimental evidence suggests that the enzymes of this pathway form a metabolon. Homology modeling of the three membrane bound proteins was carried out using the Sybyl software and available relevant crystal structures. Residues involved in tight positioning of the substrates and intermediates in the active sites of CYP79A1 and CYP71E1 were identified. In both CYPs, hydrophobic surface domains close to the N-terminal trans-membrane anchor and between the F' and G helices were identified as involved in membrane anchoring. The proximal surface of both CYPs showed positively charged patches complementary to a negatively charged bulge on CPR carrying the FMN domain. A patch of surface exposed, positively charged amino acid residues positioned on the opposite face of the membrane anchor was identified in CYP71E1 and might be involved in binding UGT85B1 via a hypervariable negatively charged loop in this protein.
authors.affiliation: Plant Biochemistry Laboratory, Department of Plant Biology and Biotechnology, University of Copenhagen, 40 Thorvaldsensvej, DK-1871 Frederiksberg C, Copenhagen, Denmark.
authors.forename: Kenneth
authors.initials: K
authors.lastname: Jensen
authors.forename: Sarah Anne
authors.initials: SA
authors.lastname: Osmani
authors.forename: Thomas
authors.initials: T
authors.lastname: Hamann
authors.forename: Peter
authors.initials: P
authors.lastname: Naur
authors.forename: Birger Lindberg
authors.initials: BL
authors.lastname: Møller
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D001665:Binding Sites; D020134:Catalytic Domain; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000378:metabolism*; D005964:Glucosyltransferases / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000378:metabolism; D009251:NADPH-Ferrihemoprotein Reductase / Q000378:metabolism; D009570:Nitriles / Q000378:metabolism*; D010940:Plant Proteins / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D012984:Software; D045868:Sorghum / Q000737:chemistry* / Q000378:metabolism; D013379:Substrate Specificity; D013499:Surface Properties; D014443:Tyrosine / Q000378:metabolism
chemical_list: D000596:Amino Acids; D008565:Membrane Proteins; D009097:Multienzyme Complexes; D009570:Nitriles; D010940:Plant Proteins; D014443:Tyrosine; D003577:Cytochrome P-450 Enzyme System; C089817:cytochrome P450TYR; C110889:cytochrome P-450 CYP71E1 (sorghum); D009251:NADPH-Ferrihemoprotein Reductase; D005964:Glucosyltransferases; C011220:dhurrin