Organization of the multifunctional enzyme type 1: interaction between N- and C-terminal domains is required for the hydratase-1/isomerase activity

Abstract

Rat peroxisomal multifunctional enzyme type 1 (perMFE-1) is a monomeric protein of beta-oxidation. We have defined five functional domains (A, B, C, D and E) in the perMFE-1 based on comparison of the amino acid sequence with homologous proteins from databases and structural data of the hydratase-1/isomerases (H1/I) and (3 S )-hydroxyacyl-CoA dehydrogenases (HAD). Domain A (residues 1-190) comprises the H1/I fold and catalyses both 2-enoyl-CoA hydratase-1 and Delta(3)-Delta(2)-enoyl-CoA isomerase reactions. Domain B (residues 191-280) links domain A to the (3 S )-dehydrogenase region, which includes both domain C (residues 281-474) and domain D (residues 480-583). Domains C and D carry features of the dinucleotide-binding and the dimerization domains of monofunctional HADs respectively. Domain E (residues 584-722) has sequence similarity to domain D of the perMFE-1, which suggests that it has evolved via partial gene duplication. Experiments with engineered perMFE-1 variants demonstrate that the H1/I competence of domain A requires stabilizing interactions with domains D and E. The variant His-perMFE (residues 288-479)Delta, in which the domain C is deleted, is stable and has hydratase-1 activity. It is proposed that the extreme C-terminal domain E in perMFE-1 serves the following three functions: (i) participation in the folding of the N-terminus into a functionally competent H1/I fold, (ii) stabilization of the dehydrogenation domains by interaction with the domain D and (iii) the targeting of the perMFE-1 to peroxisomes via its C-terminal tripeptide.

Cloning and expression pattern of peroxisomal β-oxidation genes palmitoyl-CoA oxidase, multifunctional protein and 3-ketoacyl-CoA thiolase in mussel Mytilus galloprovincialis and thicklip grey mullet Chelon labrosus

“…1), coding for the (3S)-dehydrogenase region (Kiema et al, 2002). MFP2 plays a central role in peroxisomal β-oxidation as it handles most peroxisomal β-oxidation substrates (Huyghe et al, 2006), those derived from AOX2 and AOX3 oxidation steps and the enoyl-CoAs of VLCFAs oxidised by AOX1.…”

Biochemistry and genetics of inherited disorders of peroxisomal fatty acid metabolism

“…The latter one consists of two subdomains, one (amino acids 281-474) containing a Rossman fold binding the dinucleotide with a G 304 XGXXG motif, the other one (amino acids 480-583) displaying homology to the dimerization domain of mitochondrial acyl-CoA dehydrogenases. The last domain carries the PTS1 and might be involved in protein folding and stabilizing the two active domains ( 111 ( 112 ).…”

Crystal Structure of Liganded Rat Peroxisomal Multifunctional Enzyme Type 1

“…Sequence analyses have shown that the N-and C-terminal parts belong to two different superfamilies. The N-terminal part belongs to the crotonase superfamily (4), and the C-terminal part belongs to the HAD superfamily (5).…”

“…Domains D and E are tightly associated. The evolutionary relationship of the D-and E-domains has been described previously (1,5).…”