Mitochondrial and peroxisomal targeting of 2-methylacyl-CoA racemase in humans.
2-Methylacyl-CoA racemase is an auxiliary enzyme required for the peroxisomal beta-oxidative breakdown of (2R)-pristanic acid and the (25R)-isomer of C(27) bile acid intermediates. The enzyme activity is found not only in peroxisomes but also is present in mitochondria of human liver and fibroblasts. The C terminus of the human racemase, a protein of 382 amino acids with a molecular mass of 43,304 daltons as deduced from its cloned cDNA, consists of KASL. Hitherto this sequence has not been recognized as a peroxisomal targeting signal (PTS1). From the in vitro interaction between recombinant racemase and recombinant human PTS1 receptor (Pex5p), and the peroxisomal localization of green fluorescent protein (GFP) fused to the N terminus of full-length racemase or its last six amino acids in tranfected Chinese hamster ovary (CHO) cells, we concluded that ASL is a new PTS1 variant. To be recognized by Pex5p, however, the preceding lysine residue is critical. As shown in another series of transfection experiments with GFP fused to the C terminus of the full-length racemase or racemase with deletions of the N terminus, mitochondrial targeting information is localized between amino acids 22 and 85.Hence, our data show that a single transcript gives rise to a racemase protein containing two topogenic signals, explaining the dual cellular localization of the activity.
Amery
L
L
Katholieke Universiteit Leuven, Campus Gasthuisberg, Departement Moleculaire Celbiologie, Afdeling Farmakologie, B-3000 Leuven, Belgium.
Fransen
M
M
De Nys
K
K
Mannaerts
G P
GP
Van Veldhoven
P P
PP
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D006224:Cricetinae; D049452:Green Fluorescent Proteins; D006801:Humans; D008099:Liver / Q000201:enzymology / Q000648:ultrastructure*; D008164:Luminescent Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008930:Mitochondria, Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D000074435:Peroxisome-Targeting Signal 1 Receptor; D020675:Peroxisomes / Q000201:enzymology*; D019998:Racemases and Epimerases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D017385:Sequence Homology; D014162:Transfection
D008164:Luminescent Proteins; D000074435:Peroxisome-Targeting Signal 1 Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins; D019998:Racemases and Epimerases; C108352:alpha-methylacyl-CoA racemase