Department of Biomedicine, University of Bergen, N-5020 Bergen, Norway.
Department of Biological Sciences, University of Bergen, N-5020 Bergen, Norway.
Biosci Rep. 2021 Mar 26;41(3). doi: 10.1042/BSR20202828.
N-terminal acetylation is an irreversible protein modification that primarily occurs co-translationally, and is catalyzed by a highly conserved family of N-terminal acetyltransferases (NATs). The NatC complex (NAA30-NAA35-NAA38) is a major NAT enzyme, which was first described in yeast and estimated to N-terminally acetylate ∼20% of the proteome. The activity of NatC is crucial for the correct functioning of its substrates, which include translocation to the Golgi apparatus, the inner nuclear membrane as well as proper mitochondrial function. We show in comparative viability and growth assays that yeast cells lacking MAK3/NAA30 grow poorly in non-fermentable carbon sources and other stress conditions. By using two different experimental approaches and two yeast strains, we show that liquid growth assays are the method of choice when analyzing subtle growth defects, keeping loss of information to a minimum. We further demonstrate that human NAA30 can functionally replace yeast MAK3/NAA30. However, this depends on the genetic background of the yeast strain. These findings indicate that the function of MAK3/NAA30 is evolutionarily conserved from yeast to human. Our yeast system provides a powerful approach to study potential human NAA30 variants using a high-throughput liquid growth assay with various stress conditions.
N-端乙酰化是一种不可逆的蛋白质修饰，主要发生在共翻译过程中，由高度保守的 N-端乙酰转移酶（NATs）家族催化。NatC 复合物（NAA30-NAA35-NAA38）是一种主要的 NAT 酶，最初在酵母中描述，估计可以 N-端乙酰化约 20%的蛋白质组。NatC 的活性对其底物的正常功能至关重要，这些底物包括向高尔基体、内核膜以及适当的线粒体功能的易位。我们在比较生存力和生长测定中表明，缺乏 MAK3/NAA30 的酵母细胞在非发酵碳源和其他应激条件下生长不良。通过使用两种不同的实验方法和两种酵母菌株，我们表明，当分析细微的生长缺陷时，液体生长测定是首选方法，可以最大限度地减少信息丢失。我们进一步证明，人 NAA30 可以替代酵母 MAK3/NAA30 的功能。然而，这取决于酵母菌株的遗传背景。这些发现表明，从酵母到人，MAK3/NAA30 的功能是进化保守的。我们的酵母系统提供了一种强大的方法，可使用各种应激条件的高通量液体生长测定来研究潜在的人类 NAA30 变体。
