Graduate School of Medicine, Yamaguchi University, Ube, 755-8505, Japan.
Department of Agroindustrial Technology, Faculty of Agricultural Technology, Brawijaya University, Malang, 65145, Indonesia.
Appl Microbiol Biotechnol. 2019 Jan;103(1):395-410. doi: 10.1007/s00253-018-9462-y. Epub 2018 Nov 5.
To analyze the glucose repression mechanism in the thermotolerant yeast Kluyveromyces marxianus, disrupted mutants of genes for Mig1 and Rag5 as orthologs of Mig1 and Hxk2, respectively, in Saccharomyces cerevisiae were constructed, and their characteristics were compared with those of the corresponding mutants of S. cerevisiae. MIG1 mutants of both yeasts exhibited more resistance than the corresponding parental strains to 2-deoxyglucose (2-DOG). Histidine was found to be essential for the growth of Kmmig1, but not that of Kmrag5, suggesting that MIG1 is required for histidine biosynthesis in K. marxianus. Moreover, Kmrag5 and Schxk2 were more resistant than the corresponding MIG1 mutant to 2-DOG, and only the latter increased the utilization speed of sucrose in the presence of glucose. Kmrag5 exhibited very low activities for gluco-hexokinase and hexokinase and, unlike Schxk2, showed very slow growth and a low level of ethanol production in a glucose medium. Furthermore, Kmrag5, but not Kmmig1, exhibited high inulinase activity in a glucose medium and exhibited greatly delayed utilization of accumulated fructose in the medium containing both glucose and sucrose. Transcription analysis revealed that the expression levels of INU1 for inulinase and GLK1 for glucokinase in Kmrag5 were higher than those in the parental strain; the expression level of INU1 in Kmmig1 was higher, but the expression levels of RAG1 for a low-affinity glucose transporter in Kmmig1 and Kmrag5 were lower. These findings suggest that except for regulation of histidine biosynthesis, Mig1 and Rag5 of K. marxianus play similar roles in the regulation of gene expression and share some functions with Mig1 and Hxk2, respectively, in S. cerevisiae.
为了分析产朊假丝酵母 Kluyveromyces marxianus 中的葡萄糖阻遏机制，构建了酿酒酵母 Mig1 和 Rag5 的同源基因的缺失突变体，分别与酿酒酵母中的 Mig1 和 Hxk2 进行了比较，并比较了它们与相应的酿酒酵母突变体的特征。两种酵母的 MIG1 突变体对 2-脱氧葡萄糖（2-DOG）的抗性均高于相应的亲本菌株。发现组氨酸对于 Kmmig1 的生长是必需的，但 Kmrag5 却不是，这表明 MIG1 是产朊假丝酵母中组氨酸生物合成所必需的。此外，Kmrag5 和 Schxk2 比相应的 MIG1 突变体更能抵抗 2-DOG，并且只有后者在存在葡萄糖的情况下增加了蔗糖的利用速度。Kmrag5 表现出非常低的葡糖-己糖激酶和己糖激酶活性，与 Schxk2 不同，在葡萄糖培养基中生长缓慢，乙醇产量低。此外，Kmrag5 而不是 Kmmig1 在葡萄糖培养基中表现出高的菊粉酶活性，并且在含有葡萄糖和蔗糖的培养基中，Kmrag5 表现出对积累的果糖的利用延迟。转录分析表明，Kmrag5 中菊粉酶的 INU1 和葡萄糖激酶的 GLK1 的表达水平高于亲本菌株；Kmmig1 中的 INU1 表达水平更高，但 Kmmig1 和 Kmrag5 中的 Rag1 （低亲和力葡萄糖转运蛋白）的表达水平较低。这些发现表明，除了对组氨酸生物合成的调节外，产朊假丝酵母的 Mig1 和 Rag5 在基因表达的调节中也起着相似的作用，并分别与酿酒酵母中的 Mig1 和 Hxk2 具有一些功能。
