Lecithin retinol acyltransferase contains cysteine residues essential for catalysis.

Abstract

Lecithin retinol acyltransferase (LRAT) is an essential enzyme in vitamin A metabolism and mobilization. The membrane-bound enzyme catalyzes the transfer of an acyl group from the sn-1 position of lecithin to vitamin A to generate retinyl esters. The sequence of LRAT is novel and hence does not suggest a mechanistic class to which the enzyme belongs. However, the activity of the enzyme is exceedingly sensitive to affinity labeling and group-specific reagents directed toward thiol groups. LRAT from human retinal pigment epithelium has cysteine residues at positions 161, 168, 182, and 208. Site-specific mutagenic studies show that C182 and C208 can be converted to alanines with little affect on activity. The activities of the C161A and C168A mutants are virtually nil. Moreover, while C168S is substantially active, C161S possesses only a few percent of the activity of wild-type (WT) LRAT. Also, pH-rate profiles show that C168S has virtually the same profile as WT LRAT, while C161S shows an aberrant profile quite unlike that of WT LRAT. Therefore, LRAT is a thiol acyltransferase and C161 may be the essential nucleophilic residue critical for catalysis.