title: The signal-anchor sequence of CYP2C1 inserts into the membrane as a hairpin structure.
abstract: Microsomal cytochrome P450s (CYPs) are anchored to the endoplasmic reticulum membrane by the N-terminal signal-anchor sequence which is predicted to insert into the membrane as a type 1 transmembrane helix with a luminally located N-terminus. We have mapped amino acids of the CYP2C1 signal-anchor, fused to Cys-free glutathione S-transferase, within the membrane by Cys-specific labeling with membrane-impermeant maleimide polyethylene glycol. At the C-terminal end of the signal-anchor, Trp-20 was mapped to the membrane-cytosol interface and Leu-19 was within the membrane. Unexpectedly, at the N-terminal end, Glu-2 and Pro-3 were mapped to the cytoplasmic side of the membrane rather than the luminal side as expected of a type 1 transmembrane helix. Similar results were observed for the N-terminal amino acids of the signal-anchor sequences of CYP3A4 and CYP2E1. These observations indicate that contrary to the current model of the signal-anchor of CYPs as a type 1 transmembrane helix, CYP2C1, CYP2E1, and CYP3A4 are monotopic membrane proteins with N-terminal signal-anchors that have a hairpin or wedge orientation in the membrane.
authors: Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, Urbana, IL 61821, United States.
authors: Elzbieta
authors: E
authors: Szczesna-Skorupa
authors: Byron
authors: B
authors: Kemper
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002462:Cell Membrane / Q000201:enzymology; D002522:Chlorocebus aethiops; D003545:Cysteine / Q000737:chemistry; D019392:Cytochrome P-450 CYP2E1 / Q000737:chemistry / Q000235:genetics; D051544:Cytochrome P-450 CYP3A / Q000737:chemistry / Q000235:genetics; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D008301:Maleimides / Q000737:chemistry; D008969:Molecular Sequence Data; D011092:Polyethylene Glycols / Q000737:chemistry; D021382:Protein Sorting Signals; D017433:Protein Structure, Secondary
chemical_list: D008301:Maleimides; D021382:Protein Sorting Signals; C400322:cytochrome P-450 CYP2C subfamily; C043592:maleimide; D011092:Polyethylene Glycols; D003577:Cytochrome P-450 Enzyme System; D019392:Cytochrome P-450 CYP2E1; D051544:Cytochrome P-450 CYP3A; D003545:Cysteine