Roles of cysteine 161 and tyrosine 154 in the lecithin-retinol acyltransferase mechanism.
Lecithin-retinol acyltransferase (LRAT) catalyzes the transfer of an acyl moiety from the sn-1 position of lecithin to vitamin A, generating all-trans-retinyl esters. LRAT is a unique enzyme and is the founder member of an expanding group of proteins of largely unknown function. In an effort to understand the mechanism of LRAT action, it was of interest to assign the amino acid residues responsible for the two pK(a) values of 8.22 and 9.95 observed in the pH vs rate profile. Titrating C161 of LRAT with a specific affinity labeling agent at varying pH values shows that this residue has a pK(a) = 8.03. Coupled with previous studies, this titration reveals the catalytically essential C161 as the residue responsible for the ascending limb of the pH vs rate profile. Site-specific mutagenic experiments on the lysine and tyrosine residues of LRAT reveal that only the highly conserved tyrosine 154 is essential for catalytic activity. This residue is likely to be responsible for the pK(a) = 9.95 found in the pH vs rate profile. Thus, LRAT has three essential residues (C161, Y154, and H60), all of which are conserved in the LRAT family of enzymes.
Xue
Linlong
L
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 45 Shattuck Street, Boston, Massachusetts 02115, USA.
Rando
Robert R
RR
D000217:Acyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D003545:Cysteine / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008239:Lysine / Q000378:metabolism; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D016415:Sequence Alignment; D014443:Tyrosine / Q000378:metabolism*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D014764:Viral Proteins; D014443:Tyrosine; D000217:Acyltransferases; C056942:lecithin-retinol acyltransferase; D008239:Lysine; D003545:Cysteine