The acetyl co-enzyme A synthetase genes of Kluyveromyces lactis
The acetyl co-enzyme A synthetase genes of Kluyveromyces lactis
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DOI：
10.1002/yea.936
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发表时间：
2003-01-15
期刊：
影响因子：
2.6
通讯作者：
Steensma, HY
中科院分区：
文献类型：
--
作者：
Zeeman, AM;Steensma, HY
分离到了两个编码乙酰辅酶A合成酶同工酶的乳酸克鲁维酵母基因。我们将其中一个命名为KlACS1，因为它与酿酒酵母的ACS1基因有很高的相似性。另一个基因KlACS2与酿酒葡萄球菌ACS2的相似性大于与KlACS1或ScACS1的相似性。这表明两种等基因的分化发生在物种进化分离之前，不同的功能被保存了下来。与这个观点一致的是基因转录的调控。ScACS1和KlACS1以及ScACS2和KlACS2之间似乎维持着这种调节模式。在葡萄糖培养的细胞中不存在KlACS1转录物，而在乙醇和醋酸盐培养的细胞中转录水平很高。破坏KlACS1基因不会导致对葡萄糖或乙醇的生长缺陷。然而，在醋酸盐上的生长速率降低了两倍。KlACS2在葡萄糖和醋酸盐条件下的表达量相近，而在乙醇条件下表达量略高。该基因的零突变体在所有三种碳源上的生长速率都降低了。综上所述，这些数据表明KlACS2在葡萄糖上的生长过程中被使用，而KlACS1在醋酸上的生长过程中最占优势。两个ACS基因缺失的菌株只有在含有ACS2基因的质粒存在时才能恢复，这表明双突变体是致命的。四分体分析证实，具有推断的Klacs1Klacs2基因型的无活孢子萌发，但不能进一步分裂。因此，与酿酒酵母一样，由丙酮酸脱羧酶、乙醛脱氢酶和乙酰辅酶A合成酶形成的丙酮酸脱氢酶旁路对生长至关重要。这些结果与一株具有唯一结构丙酮酸脱羧酶基因断裂的菌株在葡萄糖上生长的结果明显矛盾。然而，残留的酶活性可能解释了这种差异，或者Acs完成了一个额外的未知功能，与它的酶活性分开。KlACS1和KlACS2序列已分别存入EMBL数据库，登录号为AF061265和AF134491。版权所有:John Wiley Sons, Ltd。
Two Kluyveromyces lactis genes encoding acetyl co-enzyme A synthetase isoenzymes were isolated. One we named KlACS1, as it has high similarity to the ACS1 gene of Saccharomyces cerevisiae. The other gene, KlACS2, showed more similarity to S. cerevisiae ACS2 than to KlACS1 or ScACS1. This suggests that divergence of the two isogenes occurred before the evolutionary separation of the species and that the different functions have been conserved. In line with this idea is the regulation of transcription of the genes. The mode of regulation appeared to be maintained between ScACS1 and KlACS1 and between ScACS2 and KlACS2. The KlACS1 transcript was absent in glucose-grown cells, whereas transcription levels in ethanol- and acetate-grown cells were high. Disruption of the KlACS1 gene did not result in growth defects on glucose or ethanol. The growth rate on acetate, however, was reduced by a factor of two. KlACS2 was expressed at similar levels during growth on glucose and acetate, whereas expression on ethanol was slightly higher. A null mutant in this gene showed a reduced growth rate on all three carbon sources. Taken together, these data suggest that KlACS2 is used during growth on glucose and that KlACS1 is most dominant during growth on acetate. Strains in which both ACS genes are deleted could only be retrieved when a plasmid containing the ACS2 gene was present, suggesting that the double mutant is lethal. Tetrad analysis confirmed that non-viable spores with a deduced Klacs1Klacs2 genotype germinated but could not divide further. It therefore appears that, as in S. cerevisiae, the pyruvate dehydrogenase bypass formed by the enzymes pyruvate decarboxylase, acetaldehyde dehydrogenase and acetyl co-enzyme A synthetase is essential for growth. These results are in apparent contradiction with the growth on glucose of a strain with a disruption in the only structural pyruvate decarboxylase gene of K. lactis. Residual enzyme activity might, however, account for this discrepancy, or Acs fulfils an additional as yet unknown function, separate from its enzymatic activity. The sequences of KlACS1 and KlACS2 have been deposited in the EMBL database under Accession Nos AF061265 and AF134491, respectively. Copyright (C) 2002 John Wiley Sons, Ltd.