Aromatic boronic acids as probes of the catalytic site of human plasma lecithin-cholesterol acyltransferase

Biochim Biophys Acta. 1987 Apr 3;918(2):175-88. doi: 10.1016/0005-2760(87)90193-7.


We have recently proposed a catalytic mechanism for human plasma lecithin-cholesterol acyltransferase (EC (J. Biol. Chem. (1986) 261, 7032-7043), implicating single serine and histidine residues in phosphatidylcholine cleavage and two cysteine residues in cholesterol esterification. We now confirm the involvement of serine and histidine in catalysing the phospholipase A2 action of lecithin-cholesterol acyltransferase by demonstrating the inhibition of this activity by phenylboronic acid (Ki = 1.23 mM) and m-aminophenylboronic acid (Ki = 2.32 mM), inhibitors of known serine/histidine hydrolases. The specificity of the interaction of aromatic boronic acids with catalytic serine and histidine residues and the putative formation of a tetrahedral adduct between boron and the lecithin-cholesterol acyltransferase serine hydroxyl group which is similar to the transition-state intermediate formed between phosphatidylcholine and the catalytic serine residue was suggested by: substrate protection against inhibition by phenylboronic acids; a much reduced incorporation of phenylmethane[35S]sulphonyl fluoride into the enzyme in the presence of phenylboronic acid; the lack of interaction of histidine- or serine-modified enzyme with immobilized phenylboronic acid in the presence of glycerol (Ve/Vo = 2.7 and 2.3 respectively) when compared to the native enzyme (Ve/Vo = 5.25). Fatty acyl-lecithin-cholesterol acyltransferase, produced by incubation of the enzyme with a lecithin-apolipoprotein A-I proteoliposome substrate, was not retarded upon the sorbent column (Ve/Vo = 1.5). Modification of the enzyme's two free cysteine residues with 5,5'-dithiobis(2-nitrobenzoic acid) or potassium ferricyanide reduced (Ve/Vo = 3.5) but did not abolish retardation on the sorbent column, indicating that these modifications resulted in steric hinderance of the interaction of the boron atom with the lecithin-cholesterol acyltransferase serine hydroxyl group. These data suggest that the serine and histidine residues are proximal within the enzyme catalytic site and that both cysteine thiol groups are close to the serine hydroxyl group. The presence of significant amino-acid sequence homologies between lecithin-cholesterol acyltransferase, triacylglycerol lipases and the transacylases of fatty acid synthase is also reported.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites / drug effects
  • Boronic Acids / pharmacology*
  • Catalysis
  • Chromatography / methods
  • Cysteine
  • Histidine
  • Humans
  • Hydrogen-Ion Concentration
  • Kinetics
  • Phenylmethylsulfonyl Fluoride / metabolism
  • Phosphatidylcholine-Sterol O-Acyltransferase / antagonists & inhibitors
  • Phosphatidylcholine-Sterol O-Acyltransferase / blood*
  • Serine


  • Boronic Acids
  • Serine
  • Histidine
  • Phenylmethylsulfonyl Fluoride
  • Phosphatidylcholine-Sterol O-Acyltransferase
  • Cysteine
  • benzeneboronic acid