High‐affinity binding of fatty acyl‐CoAs and peroxisome proliferator‐CoA esters to glutathione S‐transferases

Abstract

Acyl-CoAs are present at high concentrations within the cell, yet are strongly buffered by specific binding proteins in order to maintain a low intracellular unbound acyl-CoA concentration, compatible with their metabolic role, their importance in cell signaling, and as protection from their detergent properties. This intracellular regulation may be disrupted by nonmetabolizables acyl-CoA esters of xenobiotics, such as peroxisome proliferators, which are formed at relatively high concentration within the liver cell. The low molecular mass acyl-CoA binding protein (ACBP) and fatty acyl-CoA binding protein (FABP) have been proposed as the buffering system for fatty acyl-CoAs. Whether these proteins also bind xenobiotic-CoA is not known. Here we have identified new liver cytosolic fatty acyl-CoA and xenobiotic-CoA binding sites as glutathione S-transferase (GST), using fluorescent polarization and a acyl-etheno-CoA derivative of the peroxisome proliferator nafenopin 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 xenobiotic-CoAs and fatty acyl-CoAs, with K d values ranging from 200 nm to 5 mm. One mol of acyl-CoA is bound per mol of dimeric enzyme, and no metabolization or hydrolysis was observed. Binding results in strong inhibition of rat liver GST and human recombinant GSTA1-1 (IC 50 at the nanomolar level 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 domain. Results suggest that under increased acyl-CoA concentration, as occurs after exposure to peroxisome proliferators, acyl-CoA binding to the abundant class alpha GSTs may result in strong inhibition of xenobiotic detoxification. Analysis of the binding properties of GSTs and other acyl-CoA binding proteins suggest that under increased acyl-CoA concentration GSTs would be responsible for xenobiotic-CoA binding whereas ACBP would preferentially bind fatty acyl-CoAs.

Interaction of the Electrophilic Ketoprofenyl-Glucuronide and Ketoprofenyl-Coenzyme A Conjugates with Cytosolic Glutathione S-Transferases

“…Various proteins such as albumin (Li et al, 2003b), sulfotransferases (Tulik et al, 2002), COX (Levoin et al, 2004), hepatocyte nuclear factor-4␣ (Hertz et al, 2001), and glucose-6-phosphate dehydrogenase (Asensio et al, 2007) have indeed been reported to interact with CoA thioester derivatives. It was found previously that long-chain saturated fatty acyl-CoAs and peroxisome proliferator-CoA thioesters exert high affinity toward GST (Silva et al, 1999). Our results provide another example of the ability of carboxylic acid-containing compound to bind to GST.…”

“…Furthermore, xenobiotic-SCoA metabolites were found to be much more reactive toward GSH than acylglucuronides (Olsen et al, 2002). Whereas Silva et al (1999) highlighted the absence of metabolism and hydrolysis of the thioester derivatives, interestingly, a transacylation reaction with GSH, enhanced in the presence of GST, was detected in the present study. MALDI-TOF MS analysis of KPF-SG obtained by enzymatic synthesis led to the identification of the expected ion MH ϩ m/z 544 (Fig.…”

“…A study of the interaction of KPF-OG and KPF-SCoA with cytosolic GST was undertaken. The choice of these enzymes as putative targets for these metabolites was dictated by previous reports showing that 1) various electrophilic esters can be substrates for GST (Hayes et al, 2005), including acylglucuronides (Shore et al, 1995) and acyl-CoA derivatives , 2) fatty acyl-CoA esters are potent inhibitors of recombinant GST (Silva et al, 1999), and 3) several CoA esters of xenobiotics can react with reduced glutathione (GSH), the cosubstrate of GST, leading to the corresponding thioester derivatives (references therein). The inhibition potency of KPF-OG and KPFSCoA toward cytosolic rat liver GST was evaluated and the reactivity of KPF-SCoA in the presence of these enzymes was elucidated and kinetically characterized in the present work.…”

Lipid sensing and lipid sensors

“…There has been some recent evidence that PPARs can bind to and are activated by fatty acid-CoA esters [79,80]. Conversely, it has been shown that L-FABP also binds different fatty acid-CoA esters, albeit with lower affinity than fatty acids [53,81,82]. Furthermore, it was shown that overexpression of L-FABP in fibroblasts led to 2472 C. Wolfrum Intracellular fatty acid sensing increased targeting of a fluorescently labeled fatty acid-CoA ester or free fatty acids into the nuclear membrane and nucleoplasm [82], where it colocalized with PPARa.…”