During alcohol fermentation, Saccharomyces cerevisiae produces organic acids, including succinate, acetate, and malate. Since malate contributes to the pleasant flavor of sake (a Japanese alcoholic beverage), various methods for breeding high-malate-producing yeast have been developed. We previously isolated a high-malate-producing strain and found that a missense mutation in GID4 was responsible for the high-malate-producing phenotype. Gid4 is a component of the GID (glucose-induced degradation-deficient) complex and stimulates the catabolic degradation of gluconeogenic enzymes. In this study, the mechanism by which this mutation led to high malate production in yeast cells was investigated. The evaluation of disruptants and mutants of gluconeogenic enzymes revealed that cytosolic malate dehydrogenase (Mdh2) participated in the malate production. Furthermore, target proteome analysis indicated that an increase in malate production resulted from the accumulation of Mdh2 in gid4 disruptant due to the loss of GID complex-mediated degradation. Next, we investigated the effects of GID protein-coding genes (GID1-GID9) on organic acid production and enzyme expression profiles in yeast. The disruptants of GID1, 2, 3, 4, 5, 8, and 9 exhibited high malate production. Comparison of protein abundance among the GID disruptants revealed variations in protein expression profiles, including in glycolysis and tricarboxylic acid cycle-related enzymes. The high-malate-producing disruptants showed the activation of several glycolytic enzymes and a reduction in enzymes involved in the conversion of pyruvate to ethanol. Our results suggest that high-malate-producing disruptants adapt their metabolism to produce malate in excess via the regulation of protein expression in glucose assimilation and ethanol fermentation. KEY POINTS: An increase in malate level of GID4 mutant resulted from the accumulation of Mdh2. The disruptants of GID1, 2, 3, 4, 5, 8, and 9 showed high malate production. The protein expression profiles in the GID disruptants differed from one another.

中文翻译：

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GID蛋白编码基因突变对酿酒酵母中苹果酸产生和酶表达谱的影响。

在酒精发酵过程中，酿酒酵母会产生有机酸，包括琥珀酸，乙酸盐和苹果酸。由于苹果酸有助于清酒（日本酒精饮料）的令人愉悦的风味，因此已经开发了多种繁殖高苹果酸的酵母的方法。我们先前分离出了高苹果酸生成菌株，并发现GID4的错义突变是造成高苹果酸生成表型的原因。Gid4是GID（葡萄糖诱导的降解缺陷）复合物的组成部分，可刺激糖异生酶的分解代谢降解。在这项研究中，研究了这种突变导致酵母细胞中高苹果酸产生的机制。对糖异生酶的破坏剂和突变体的评估表明，胞质苹果酸脱氢酶（Mdh2）参与了苹果酸的生产。此外，目标蛋白质组分析表明，由于GID复合物介导的降解损失，Mid2在gid4破坏物中的积累导致了苹果酸产量的增加。接下来，我们研究了GID蛋白编码基因（GID1-GID9）对酵母中有机酸产生和酶表达谱的影响。GID1、2、3、4、5、8和9的破坏物表现出高苹果酸产量。GID破坏剂之间的蛋白质丰度比较表明，蛋白质表达谱存在变化，包括糖酵解和三羧酸循环相关的酶。产生高苹果酸的破坏剂显示出几种糖酵解酶的活化和参与丙酮酸向乙醇转化的酶的减少。我们的结果表明，产生高苹果酸的破坏剂通过调节葡萄糖同化和乙醇发酵中的蛋白质表达来调节其代谢，从而过量生产苹果酸。要点：GID4突变体的苹果酸水平升高是由于Mdh2的积累引起的。GID1、2、3、4、5、8和9的破坏物显示出高的苹果酸产量。GID破坏剂中的蛋白质表达谱互不相同。