Molecular cloning of cDNA species for rat and mouse liver alpha-methylacyl-CoA racemases.
cDNA species coding for alpha-methylacyl-CoA racemase were cloned from rat and mouse liver cDNA libraries and characterized. The rat liver lambdagt11 cDNA expression library was screened with anti-racemase IgG [Schmitz, Albers, Fingerhut and Conzelmann (1995) Eur. J. Biochem.231, 815-822]. Several full-length clones were obtained that contained an open reading frame of 1083 bp, coding for a protein of 361 amino acid residues with a predicted molecular mass of 39679 Da. The sequences of three peptides that were isolated by HPLC from a tryptic digest of purified rat liver racemase fully matched the cDNA-derived amino acid sequence. The cDNA coding for mouse racemase was cloned from a mouse liver lambdaZAP cDNA expression library and sequenced. The coding region of 1080 bp codes for a 360-residue protein (molecular mass 39558 Da) that shares 89.7% similarity with the rat protein. Expression of the rat racemase as are combinant protein in Escherichia coli with the pTrcHisB-expression vector yielded enzymically active protein. The amino acid sequences of alpha-methylacyl-CoA racemases do not resemble any known sequence of beta-oxidation or auxiliary enzymes, supporting the view of a highly diverse evolutionary origin of enzymes acting on fatty acyl-CoA S-esters.
Schmitz
W
W
Theodor-Boveri-Institut für Biowissenschaften (Biozentrum) der Universität Würzburg, Am Hubland, D-97074 Würzburg, Federal Republic of Germany.
Helander
H M
HM
Hiltunen
J K
JK
Conzelmann
E
E
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D008099:Liver / Q000201:enzymology*; D051379:Mice; D008969:Molecular Sequence Data; D019998:Racemases and Epimerases / Q000235:genetics*; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics; D016415:Sequence Alignment
D018076:DNA, Complementary; D011994:Recombinant Proteins; D019998:Racemases and Epimerases; C108352:alpha-methylacyl-CoA racemase