Biochemical characterization of the PHARC associated serine hydrolase ABHD12 reveals its preference for very long chain lipids

Abstract

Polyneuropathy, hearing loss, ataxia, retinitis pigmentosa, and cataract (PHARC) is a rare genetic human neurological disorder caused by null mutations to the Abhd12 gene, which encodes the integral membrane serine hydrolase enzyme ABHD12. While the role that ABHD12 plays in PHARC is understood, the thorough biochemical characterization of ABHD12 is lacking. Here, we report the facile synthesis of mono-1-(fatty)acyl-glycerol lipids of varying chain lengths and unsaturation, and use this lipid substrate library, to biochemically characterize recombinant mammalian ABHD12. The substrate profiling study for ABHD12 suggested that this enzyme requires glycosylation for optimal activity, and that is has a strong preference for very long chain lipid substrates. We further validated this substrate profile against brain membrane lysates generated from wild type and ABHD12 knockout mice. Finally, using cellular organelle fractionation and immunofluorescence assays, we show that mammalian ABHD12 is enriched on the endoplasmic reticulum membrane, where most of the very long chain fatty acids are biosynthesized in cells. Taken together, our findings provide a biochemical explanation for why very long chain lipids (such as lysophosphatidylserine lipids) accumulate in the brains of ABHD12 knockout mice, which is a murine model of PHARC.