Secondary structure of a truncated form of lecithin retinol acyltransferase in solution and evidence for its binding and hydrolytic action in monolayers.
Lecithin retinol acyltransferase (LRAT) is a 230 amino acids membrane-associated protein which catalyzes the esterification of all-trans-retinol into all-trans-retinyl ester. The enzymatic activity of a truncated form of LRAT (tLRAT) which contains the residues required for catalysis but which is lacking N- and C-terminal hydrophobic segments has been shown to depend on the detergent used for its solubilization. Moreover, it is unknown whether tLRAT can bind membranes in the absence of these hydrophobic segments. The present study has allowed to measure the membrane binding and hydrolytic action of tLRAT in lipid monolayers by use of polarization modulation infrared reflection absorption spectroscopy and Brewster angle microscopy. Moreover, the proportion of the secondary structure components of tLRAT was determined in three different detergents by infrared absorption spectroscopy, vibrational circular dichroism and electronic circular dichroism which allowed to explain its detergent dependent activity. In addition, the secondary structure of tLRAT in the absence of detergent was very similar to that in Triton X-100 thus suggesting that, compared to the other detergents assayed, the secondary structure of this protein is very little perturbed by this detergent.
Bussières
Sylvain
S
Unité de recherche en ophtalmologie, Centre Hospitalier Universitaire de Québec, Pavillon CHUL, Département d'ophtalmologie, Faculté de médecine, Université Laval, 2705 Blvd. Laurier, Ste-Foy, Québec, Canada G1V 4G2.
Buffeteau
Thierry
T
Desbat
Bernard
B
Breton
Rock
R
Salesse
Christian
C
D000217:Acyltransferases / Q000737:chemistry* / Q000378:metabolism; D001483:Base Sequence; D002942:Circular Dichroism; D017931:DNA Primers; D003902:Detergents / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013055:Spectrophotometry, Infrared
D017931:DNA Primers; D003902:Detergents; D011994:Recombinant Proteins; D000217:Acyltransferases; C056942:lecithin-retinol acyltransferase