Studies of Human 2,4-Dienoyl CoA Reductase Shed New Light on Peroxisomal β-Oxidation of Unsaturated Fatty Acids

Tian Hua,Dong Wu,Wei Ding,Jiangyun Wang,Neil Shaw,Zhi-Jie Liu

doi:10.1074/jbc.m112.385351

Abstract

Peroxisomes play an essential role in maintaining fatty acid homeostasis. Although mitochondria are also known to participate in the catabolism of fatty acids via β-oxidation, differences exist between the peroxisomal and mitochondrial β-oxidation. Only peroxisomes, but not mitochondrion, can shorten very long chain fatty acids. Here, we describe the crystal structure of a ternary complex of peroxisomal 2,4-dienoyl CoA reductases (pDCR) with hexadienoyl CoA and NADP, as a prototype for comparison with the mitochondrial 2,4-dienoyl CoA reductase (mDCR) to shed light on the differences between the enzymes from the two organelles at the molecular level. Unexpectedly, the structure of pDCR refined to 1.84 Å resolution reveals the absence of the tyrosine-serine pair seen in the active site of mDCR, which together with a lysine and an asparagine have been deemed a hallmark of the SDR family of enzymes. Instead, aspartate hydrogen-bonded to the Cα hydroxyl via a water molecule seems to perturb the water molecule for protonation of the substrate. Our studies provide the first structural evidence for participation of water in the DCR-catalyzed reactions. Biochemical studies and structural analysis suggest that pDCRs can catalyze the shortening of six-carbon-long substrates in vitro. However, the K(m) values of pDCR for short chain acyl CoAs are at least 6-fold higher than those for substrates with 10 or more aliphatic carbons. Unlike mDCR, hinge movements permit pDCR to process very long chain polyunsaturated fatty acids.

Highlights

Reasons for the differences in activity between the peroxisomal and mitochondrial DCRs are unknown
The structure of peroxisomal DCR (pDCR) refined to 1.84 Å resolution reveals the absence of the tyrosine-serine pair seen in the active site of mDCR, which together with a lysine and an asparagine have been deemed a hallmark of the short chain dehydrogenase/reductase (SDR) family of enzymes
Mechanism of Catalysis— the conserved catalytic serine of the SDR family could not be identified by primary sequence alignment for mDCR, a structural view of mDCR confirmed the presence of Tyr-199 and Ser-210 at the catalytic

Summary

Background

Reasons for the differences in activity between the peroxisomal and mitochondrial DCRs are unknown. Results: Structure-function studies on peroxisomal DCR (pDCR) and comparison with its mitochondrial counterpart reveal differences in catalytic residues and hinge movements. In contrast to humans, ␤-oxidation in peroxisomes from yeast and plants proceeds to completion [12] The reason for this difference has been attributed to the composition of the active site: in particular, the ability of the enzyme to orient the shorter substrate correctly for catalysis [13]. 2,4-dienoyl CoA reductase produces trans-3-enoyl CoA, which is isomerized by ⌬3,⌬2-enoyl CoA isomerase to trans-2-enoyl CoA Among all of these enzymatic steps, the reaction catalyzed by 2,4-dienoyl CoA reductase is considered to be the rate-limiting step in the ␤-oxidation of polyunsaturated fatty acids [16]. Version of 2,4-dienoyl CoA to trans-3-enoyl CoA is the ratelimiting step in the catabolism of poly unsaturated fatty acids, the mechanistic insights gained in this study have profound implications for treatment of diseases characterized by accumulation of polyunsaturated fatty acids

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION