Biosynthesis of Branched-Chain Amino Acids in Yeast: Regulation of Leucine Biosynthesis in Prototrophic and Leucine Auxotrophic Strains

Abstract

The first enzyme in the biosynthesis of leucine in yeast, a-isopropylmalate synthetase, is inhibited by L-leucine. In a mutant resistant to the analogue 5',5',5'trifluoroleucine, the enzyme is markedly resistant to inhibition by L-leucine. Growth ing the presence of exogenous L-leucine results in repression of the second and third enzymes of the pathway. The first enzyme is not repressed unless both L-leucine and L-threonine are supplied in the medium. Comparison of levels of the remaining two enzymes in leucine auxotrophs grown under conditions of leucine excess and leucine limitation reveals deviations from the wild-type derepression pattern in some mutants. In some, repression of the synthetase by leucine alone was observed. In others, the repressibility of the dehydrogenase was lost. It is unlikely that these deviations were due to the same primary mutational event that caused leucine auxotrophy. No mutants were found in which an altered gene was recognized to be clearly responsible for the level of the leucine-forming enzymes.

Quality improvement and geographical indication of cachaça (Brazilian spirit) by using locally selected yeast strains

“…A pool of colonies from each distillery were screened in different carbon sources (glucose 2%, galactose 2%, maltose 2%), and according to stress resistance (high osmolarity (glucose 20%, sucrose 20%), high ethanol content (15% v/v) and high temperature (37°C)), as previously described (Vaughan-Martini and Martini 1993;Vicente et al 2006). Drug resistance (5,5 0 ,5 00 -trifluoro-DLleucine (TFL) 1 mmol l À1 and cerulenin 25 lmol l À1 ) were performed as previously described (Satyanarayana et al 1968;Bussey and Umbarger 1970;Ichikawa et al 1991). Other beneficial characteristics to cachac ßa and its production process were also tested (flocculation ability, absence of hydrogen sulphide production, high invertase activity and high production of esters and higher alcohols) (Vicente et al 2006).…”

Enhanced formation of isoamyl alcohol in Zygosaccharomyces rouxii due to elimination of feedback inhibition of α-isopropylmalate synthase

“…It was supposed that feedback inhibition of (Y-IPM synthase was diminished in the mutants so that a higher level of (w-IPM could be synthesized from a-keto isovalerate instead of isobutyl alcohol. In S. cereuisiae, the a-IPM synthase in parent was inhibited by 80% in the presence of r_-leucine at a concentration of 5 mM, but the CX-IPM synthase in mutant was not inhibited in the presence of L-leucine [2,3]. On the other hand, the activity of cr-IPM synthase in both parent and mutant was repressed by 60-70% when the cells were grown with L-leucine and L-threonine [3].…”

“…Cells were grown to the early stationary phase in YNB medium containing 0.67% yeast nitrogen base without amino acids (Difco Laboratories, Detroit, Ml), 2% glucose and supplemented with 2 mM L-leucine to obtain a higher level of a-IPM synthase activity [2]. The a-IPM synthase was not repressed but rather was consistently higher in cells grown with L-leucine, although both the isomerase and the dehydrogenase in the leucine synthetic pathway were repressed in yeast [2]. Crude extracts of each cell culture were prepared by the method of Ulm et al [9].…”

“…Tel. : +81 (262) 27 3131; Fax: +81 (262) 27 3130 glucose; and (ii) the Ehrlich mechanism that synthesizes (Y-KC from L-leucine [2]. In the absence of L-leucine, i-AmOH is mainly derived from a-KC through the L-leucine synthetic pathway [3].…”

“…In the absence of L-leucine, i-AmOH is mainly derived from a-KC through the L-leucine synthetic pathway [3]. On the other hand, in the presence of L-leucine, a major route of i-AmOH synthesis is thought be through the Ehrlich mechanism, since cw-isopropylmalate ( cy-IPM) synthase, the enzyme used for &-KC synthesis from glucose, is inhibited by L-leucine [2]. We supposed that a-KC could be synthesized directly from glucose and the amount of i-AmOH produced in miso fermentation would be increased under the condition that this inhibition of CY-IPM synthase was relieved.…”