Construction of a brewer's yeast having alpha-acetolactate decarboxylase gene from Acetobacter aceti ssp. xylinum integrated in the genome.

alpha-Acetolactate decarboxylase (ALDC) gene from Acetobacter aceti ssp. xylinum has several possible initiation codons in the N-terminus. To determine the initiation codon of the ALDC giving the highest expression levels, glyceraldehyde-3-phosphate dehydrogenase (GPD) promoter was linked just upstream of each possible initiation codon. The ALDC whose translation starts 130 bp downstream from the first ATG codon had the highest activity in yeast cells. When expression levels of the ALDC gene were compared using three strong yeast promoters of glycolytic genes, alcohol dehydrogenase I (ADC1), phosphoglycerate kinase (PGK) and GPD, the GPD promoter was the strongest. The ALDC gene was integrated in a ribosomal RNA gene of a brewer's yeast by co-transformation with an expression plasmid of G418-resistance gene. The laboratory-scale growth test confirmed that the total diacetyl concentration was reduced in wort.