High-affinity binding of fatty acyl-CoAs and peroxisome proliferator-CoA esters to glutathione S-transferases - Effect on enzymatic activity

Acyl-CoAs an present at high concentrations within the cell, yet are strong ly buffered by specific binding proteins in order to maintain a low intrace llular unbound acyl-CoA concentration, compatible with their metabolic role ? their importance in cell signaling, and as protection from their detergen t properties. This intracellular regulation may be disrupted by nonmetaboli zables acyl-CoA eaters of xenobiotics, such as peroxisome proliferators, wh ich are formed at relatively high concentration within the liver cell. The low molecular mass acyl-CoA binding protein (ACBP) and fatty acyl-CoA bindi ng protein (FABP) have been proposed as the buffering system for fatty acyl -CoAs. Whether these proteins also bind xenobiotic-CoA is not known. Here w e have identified new liver cytosolic fatty acyl-CoA and xenobiotic-CoA bin ding sites as glutathione S-transferase (GST), using fluorescent polarizati on and a acyl-etheno-CoA derivative of the peroxisome proliferator nafenopi n as ligand. Rat liver GST and human liver recombinant GSTA1-1, GSTP1-1 and GSTM1-1 were used. Only class alpha rat liver GST and human GSTA1-1 bind x enobiotic-CoAs and fatty acyl-CoAs, with K-d values ranging from 200 nM to 5 mu M. One mol of acyl-CoA is bound per mol of dimeric enzyme, and no meta bolization or hydrolysis was observed. Binding results in strong inhibition of rat Liver GST and human recombinant GSTA1-1 (IC50 at the nanomolar leve l for palmitoyl-CoA) but not GSTP1-1 and GSTM1-1. Acyl-CoAs do not interact with the GSTA1-1 substrate binding site, but probably with a different dom ain. Results suggest that under increased acyl-CoA concentration, as occurs after exposure to peroxisome proliferators, acyl-CoA binding to the abunda nt class alpha GSTs may result in strong inhibition of xenobiotic detoxific ation, Analysis of the binding properties of GSTs and other acyl-CoA bindin g proteins suggest that under increased acyl-CoA concentration GSTs would b e responsible for xenobiotic-CoA binding whereas ACBP would preferentially bind fatty acyl-CoAs.