Identification, cloning, and characterization of a Lactococcus lactis branched-chain alpha-keto acid decarboxylase involved in flavor formation.
The biochemical pathway for formation of branched-chain aldehydes, which are important flavor compounds derived from proteins in fermented dairy products, consists of a protease, peptidases, a transaminase, and a branched-chain alpha-keto acid decarboxylase (KdcA). The activity of the latter enzyme has been found only in a limited number of Lactococcus lactis strains. By using a random mutagenesis approach, the gene encoding KdcA in L. lactis B1157 was identified. The gene for this enzyme is highly homologous to the gene annotated ipd, which encodes a putative indole pyruvate decarboxylase, in L. lactis IL1403. Strain IL1403 does not produce KdcA, which could be explained by a 270-nucleotide deletion at the 3' terminus of the ipd gene encoding a truncated nonfunctional decarboxylase. The kdcA gene was overexpressed in L. lactis for further characterization of the decarboxylase enzyme. Of all of the potential substrates tested, the highest activity was observed with branched-chain alpha-keto acids. Moreover, the enzyme activity was hardly affected by high salinity, and optimal activity was found at pH 6.3, indicating that the enzyme might be active under cheese ripening conditions.
Smit
Bart A
BA
Department of Flavor, Nutrition and Ingredients, NIZO food research, 6710 BA Ede, The Netherlands.
van Hylckama Vlieg
Johan E T
JE
Engels
Wim J M
WJ
Meijer
Laura
L
Wouters
Jan T M
JT
Smit
Gerrit
G
D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000597:Amino Acids, Branched-Chain / Q000378:metabolism; D003001:Cloning, Molecular; D003611:Dairy Products / Q000382:microbiology; D005421:Flavoring Agents / Q000378:metabolism*; D013294:Lactococcus lactis / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity
D000597:Amino Acids, Branched-Chain; D005421:Flavoring Agents; D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)