In Mouse α-Methylacyl-CoA Racemase, the Same Gene Product Is Simultaneously Located in Mitochondria and Peroxisomes

Abstract

␣-Methylacyl-CoA racemase, an enzyme of the bile acid biosynthesis and branched chain fatty acid degradation pathway, was studied at the protein, cDNA, and genomic levels in mouse liver. Immunoelectron microscopy and subcellular fractionation located racemase to mitochondria and peroxisomes. The enzymes were purified from both organelles with immunoaffinity chromatography. The isolated proteins were of the same size, with identical N-terminal amino acid sequences, and the existence of additional proteins with ␣-methylacyl-CoA racemase activity was excluded. A racemase gene of about 15 kilobases was isolated. Southern blot analysis and chromosomal localization showed that only one racemase gene is present, on chromosome 15, region 15B1. The putative initial ATG in the racemase gene was preceded by a functional promotor as shown with the luciferase reporter gene assay. The corresponding cDNAs were isolated from rat and mouse liver. The recombinant rat protein was overexpressed in active form in Pichia pastoris. The presented data suggest that the polypeptide encoded by the racemase gene can alternatively be targeted to peroxisomes or mitochondria without modifications. It is concluded that the noncleavable N-terminal sequence of the polypeptide acts as a weak mitochondrial and that the C-terminal sequence acts as a peroxisomal targeting signal.

Role of α-Methylacyl Coenzyme A Racemase in the Degradation of Methyl-Branched Alkanes by Mycobacterium sp. Strain P101

“…Phytanic acid is derived from chlorophyll-bound phytol and is converted to pristanic acid by ␣-oxidation in the peroxisomes. The CoA thioester pristanoyl-CoA is further ␤-oxidized, first by two to three cycles in the peroxisomes and then later by cycles in the mitochondria (11,18,36).…”

Molecular Mechanisms and Physiological Significance of Organelle Interactions and Cooperation

“…For DHCA and THCA, which are produced from cholesterol in the liver, this is a potential problem because the methyl-group in the carboxy-terminal side chain of DHCA and THCA has the 2(R)-rather than 2(S)-configuration which thus prohibits beta-oxidation. This problem is resolved by an enzyme called 2-methyl-acyl-CoA racemase (AMACR), located both in peroxisomes and mitochondria, which is able to convert 2(R)-acyl-CoAs into 2(S)-acyl-CoAs (Schmitz et al, 1997; Amery Ferdinandusse et al, 2000;Kotti et al, 2000). The same enzyme is also required to convert the 2(R)-methyl-group in pristanic acid (2,10,14,16-tetramethylpentadecanoic acid) as derived from phytanic acid into the 2(S)-position ( Figure 3A).…”

Transport of substrates into peroxisomes: the paradigm of β-lactam biosynthetic intermediates

“…They are usually peroxisome-specific, although some of these proteins can have two or more subcellular localizations due to the presence of several targeting signals within the same protein, e.g., 3-hydroxymethylglutary-CoA lyase (10) and α -methylacyl-CoA racemase (11) . The β -oxidation pathway of fatty acids has a dual peroxisomal and mitochondrial localization in mammals and plants, but it is exclusively peroxisomal in yeasts (12) and filamentous fungi because mutants devoid of peroxisomes are unable to grow on oleate (13,14) .…”