Lactate and amino acid catabolism in the cheese-ripening yeast Yarrowia lipolytica

Abstract

The consumption of lactate and amino acids is very important for microbial development and/or aroma production during cheese ripening. A strain of Yarrowia lipolytica isolated from cheese was grown in a liquid medium containing lactate in the presence of a low (0.1x) or high (2x) concentration of amino acids. Our results show that there was a dramatic increase in the growth of Y. lipolytica in the medium containing a high amino acid concentration, but there was limited lactate consumption. Conversely, lactate was efficiently consumed in the medium containing a low concentration of amino acids after amino acid depletion was complete. These data suggest that the amino acids are used by Y. lipolytica as a main energy source, whereas lactate is consumed following amino acid depletion. Amino acid degradation was accompanied by ammonia production corresponding to a dramatic increase in the pH. The effect of adding amino acids to a Y. lipolytica culture grown on lactate was also investigated. Real-time quantitative PCR analyses were performed with specific primers for five genes involved in amino acid transport and catabolism, including an amino acid transporter gene (GAP1) and four aminotransferase genes (ARO8, ARO9, BAT1, and BAT2). The expression of three genes involved in lactate transport and catabolism was also studied. These genes included a lactate transporter gene (JEN1) and two lactate dehydrogenase genes (CYB2-1 and CYB2-2). Our data showed that GAP1, BAT2, BAT1, and ARO8 were maximally expressed after 15 to 30 min following addition of amino acids (BAT2 was the most highly expressed gene), while the maximum expression of JEN1, CYB2-1, and CYB2-2 was delayed (>or=60 min).

FIG. 1.

Growth (A), increase in the pH (B), and lactate consumption (C) of Y. lipolytica 1E07 in SM containing lactate and/or amino acids. AA, amino acids; OD₆₀₀, optical density at 600 nm.

FIG. 2.

Overall rates of consumption of amino acids by Y. lipolytica 1E07 cultivated in SM containing low and high concentrations of amino acids. (A) 0.1× amino acids. (B) 0.1× amino acids plus lactate. (C) 2× amino acids. (D) 2× amino acids plus lactate.

FIG. 3.

Ammonia production (bars) by Y. lipolytica 1E07 cultivated in SM containing low and high concentrations of amino acids. (A) 0.1× amino acids. (B) 0.1× amino acids plus lactate. (C) 2× amino acids. (D) 2× amino acids plus lactate. The accumulated ammonia production is indicated by filled squares.

FIG. 4.

Levels of expression of the GAP1, BAT2, BAT1, ARO8, JEN1, CYB2-1, and CYB2-2 genes, as measured by real-time PCR after an amino acid pulse (0.1× amino acids), for a Y. lipolytica 1E07 pure culture grown in SM containing lactate.

FIG. 5.

Possible relationship between amino acid catabolism (AA) and lactate catabolism in Y. lipolytica 1E07. +, early induction of the genes in italics; +/−, delayed induction of the genes following addition of amino acids. AT, amino transferase; LDH, lactate dehydrogenase; α-KG, α-ketoglutarate; Glu, glutamate. Gene functions are shown in Table 2.