L-3-hydroxyacyl coenzyme A dehydrogenase. The location of NAD binding sites and the bilobal subunit structure.

Abstract 3-Hydroxyacyl-CoA dehydrogenase (EC 1.1.1.35) is a mitochondrial enzyme involved in the beta-oxidation of long chain fatty acids. The determination of its molecular structure at 5.25-A resolution by x-ray diffraction techniques is described. Three isomorphous derivatives, K2PtCl6, methyl mercuric chloride, and IrCl3, were prepared using crystals previously soaked in an NAD-containing solution. The positions of the heavy atom sites were determined by inspection of Patterson maps and confirmed by cross-difference Fourier maps. After refinement of the heavy atom positions, electron density maps at 5.25-A resolution were calculated. Careful study of these electron density maps revealed a unique crystalline packing arrangement in which the asymmetric unit contained 1.5 dimers of L-3-hydroxyacyl coenzyme A dehydrogenase. With this packing motif, one L-3-hydroxyacyl coenzyme A dehydrogenase dimer lies in a general position in the asymmetric unit, while the other dimer is located such that its molecular 2-fold axis is coincident with a crystallographic dyad. At 5.25-A resolution, each L-3-hydroxyacyl coenzyme A dehydrogenase subunit displays a bilobal structure. The larger lobe, which binds NAD, has approximate dimensions of 37 X 45 X 35 A. The size of the smaller lobe is approximately 30 X 23 X 20 A. Difference Fourier maps between the crystalline apo- and holoenzyme have also been calculated at 5.25-A resolution, and preliminary model fitting studies show that NAD binds to L-3-hydroxyacyl coenzyme A dehydrogenase in an open conformation similar to that found in other dehydrogenases.