Biochemical and molecular characterization of alpha-ketoisovalerate decarboxylase, an enzyme involved in the formation of aldehydes from amino acids by Lactococcus lactis.
In this paper, we report for the first time on the identification, purification, and characterization of the alpha-ketoisovalerate decarboxylase from Lactococcus lactis, a novel enzyme responsible for the decarboxylation into aldehydes of alpha-keto acids derived from amino acid transamination. The kivd gene consisted of a 1647 bp open reading frame encoding a putative peptide of 61 kDa. Analysis of the deduced amino acid sequence indicated that the enzyme is a non-oxidative thiamin diphosphate (ThDP)-dependent alpha-keto acid decarboxylase included in the pyruvate decarboxylase group of enzymes. The active enzyme is a homo-tetramer that showed optimum activity at 45 degrees C and at pH 6.5 and exhibited an inhibition pattern typical for metal-dependant enzymes. In addition to Mg(2+), activity was observed in presence of other divalent cations such as Ca(2+), Co(2+) and Mn(2+). The enzyme showed the highest specific activity (80.7 Umg(-1)) for alpha-ketoisovalerate, an intermediate metabolite in valine and leucine biosynthesis. On the other side, decarboxylation of indole-3-pyruvate and pyruvate only could be detected by a 100-fold increase in the enzyme concentration present in the reaction.
de la Plaza
Marta
M
Department of Dairy Science and Technology, Instituto del Frío (CSIC), Ciudad Universitaria, José Antonio Novais 10, 28040 Madrid, Spain.
Fernández de Palencia
Pilar
P
Peláez
Carmen
C
Requena
Teresa
T
D000447:Aldehydes / Q000378:metabolism*; D000596:Amino Acids / Q000378:metabolism*; D002262:Carboxy-Lyases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D013294:Lactococcus lactis / Q000201:enzymology* / Q000235:genetics; D007930:Leucine / Q000096:biosynthesis; D016366:Open Reading Frames; D011773:Pyruvates / Q000378:metabolism; D014633:Valine / Q000096:biosynthesis
D000447:Aldehydes; D000596:Amino Acids; D011773:Pyruvates; D002262:Carboxy-Lyases; D007930:Leucine; D014633:Valine