Xenobiotic incorporation into pyruvate dehydrogenase complex can occur via the exogenous lipoylation pathway

Abstract

Lipoylated enzymes such as the E2 component of pyruvate dehydrogenase complex (PDC-E2) are targets for autoreactive immune responses in primary biliary cirrhosis, with lipoic acid itself forming a component of the dominant auto-epitopes. A candidate mechanism for the initiation of tolerance breakdown in this disease is immune recognition of neo-antigens formed by xenobiotic substitution of normal proteins. Importantly, sensitization with proteins artificially substituted with the lipoic acid analogue xenobiotic 6-bromohexanoic acid (6BH) can induce an immune response that cross-reacts with PDC-E2. This study investigated the potential of recombinant lipoylation enzymes lipoate activating enzyme and lipoyl-AMP(GMP):N-lysine lipoyl transferase to aberrantly incorporate xenobiotics into PDC-E2. It was found that these enzymes could incorporate lipoic acid analogues including octanoic and hexanoic acids and the xenobiotic 6BH into PDC-E2. The efficiency of incorporation of these analogues showed a variable dependence on activation by adenosine triphosphate (ATP) or guanosine triphosphate (GTP), with ATP favoring the incorporation of hexanoic acid and 6BH whereas GTP enhanced substitution by octanoic acid. Importantly, competition studies showed that the relative incorporation of both 6BH and lipoic acid could be regulated by the balance between ATP and GTP, with the formation of 6BH-substituted PDC-E2 predominating in an ATP-rich environment. Conclusion: Using a well-defined system in vitro we have shown that an important xenobiotic can be incorporated into PDC in place of lipoic acid by the exogenous lipoylation system; the relative levels of lipoic acid and xenobiotic incorporation may be determined by the balance between ATP and GTP. These observations suggest a clear mechanism for the generation of an autoimmunogenic neo-antigen of relevance for the pathogenesis of primary biliary cirrhosis.(HEPATOLOGY 2008;48:1874-1884.) P rimary biliary cirrhosis (PBC) is characterized by the presence of autoreactive immune responses, typically directed at members of the 2-oxo-acid dehydrogenase family of multi-enzyme complexes (2-OADC), 1,2 with specificity for components of one or more 2-OADC family members (pyruvate dehydrogenase complex [PDC], 2-oxoglutarate dehydrogenase complex [OGDC], and branchedchain oxo-acid dehydrogenase complex [BCOADC]) seen in over 95% of patients. The archetypal autoreactive response in PBC, present in over 90% of patients, is a high titre autoantibody response reactive with both the dihydrolipoamide acetyl-transferase (E2) and the dihydrolipoamide dehydrogenase (E3) binding protein (E3BP) components of PDC. 3,4 PDC-E2, PDC-E3BP and the other 2-OADC-E2 components to which immune-reactivity is seen in PBC share a highly conserved domain structure containing a lipoic acid cofactor. 2,5 This lipoic acid is bound through an amide linkage formed between its carboxylic acid group and the ⑀-amino group of a specific lysine residue and plays a fundamentally important role as a carrier of activated acyl ...

Lipoic Acid Metabolism in Microbial Pathogens

“…A second enzyme, known as lipoyl nucleoside monophosphate (NMP) transferase, then transfers lipoate from lipoyl-GMP to the apodomains (55). The mammalian LAE also functions as a xenobiotic-metabolizing/medium-chain acyl-CoA ligase (57) and can partner with the NMP transferase to aberrantly attach lipoate analogs and xenobiotics to the mammalian PDH E2 (240). Although the mammalian NMP transferases are orthologs of E. coli LplA, they are incapable of using free lipoate as substrate and present a lipoylation strategy distinct from that of E. coli and other microorganisms (55,56).…”

The accuracy of the anti-mitochondrial antibody and the M2 subtype test for diagnosis of primary biliary cirrhosis: a meta-analysis

“…Recent data suggest that it may be due to a combination of genetic [4], infectious [5], immunizing, and environmental [6] factors. It is characterized by the destruction of small, intrahepatic bile ducts, and it mainly affects women over the age of 40.…”

Infectious Agents and Xenobiotics in the Etiology of Primary Biliary Cirrhosis

“…As previously stated, lipoic acid is attached to a limited number of proteins, yet it is a critical component of the PDC-E2 epitope [80]. The PDC-E2 structure exposes lipoic acid at the exterior of the protein complex making it accessible to chemical modification [123]. The role of xenobiotics in PBC is supported by serum reactivity against specific organic compounds with structures similar to lipoic acid [124]; further, two of these compounds (6-bromohexanoate and 2-octynoic acid) are capable of inducing AMA and PBC-like liver lesions in guinea pigs [125] and NOD.1101 [126] or C57BL/6 [127] mice, respectively.…”