About 80% of human proteins are amino-terminally acetylated (Nt-acetylated) by one of seven Nt-acetyltransferases (NATs). Actin, the most abundant protein in the cytoplasm, has its own dedicated NAT, NAA80, which acts posttranslationally and affects cytoskeleton assembly and cell motility. Here, we show that NAA80 does not associate with filamentous actin in cells, and its natural substrate is the monomeric actin-profilin complex, consistent with Nt-acetylation preceding polymerization. NAA80 Nt-acetylates actin-profilin much more efficiently than actin alone, suggesting that profilin acts as a chaperone for actin Nt-acetylation. We determined crystal structures of the NAA80-actin-profilin ternary complex, representing different actin isoforms and different states of the catalytic reaction and revealing the first structure of NAT-substrate complex at atomic resolution. The structural, biochemical, and cellular analysis of mutants shows how NAA80 has evolved to specifically recognize actin among all cellular proteins while targeting all six actin isoforms, which differ the most at the amino terminus.

约80％的人类蛋白质被七个Nt-乙酰基转移酶（NATs）之一氨基末端乙酰化（Nt-乙酰化）。肌动蛋白是细胞质中最丰富的蛋白质，它有自己专用的NAT，即NAA80，它在翻译后起作用并影响细胞骨架的组装和细胞运动。在这里，我们表明，NAA80不与细胞中的丝状肌动蛋白缔合，其天然底物是单体肌动蛋白-脯氨酸蛋白复合物，与聚合前的Nt-乙酰化相一致。NAA80 Nt-乙酰化肌动蛋白-profilin比单独的肌动蛋白更有效，表明profilin充当肌动蛋白Nt-乙酰化的伴侣。我们确定了NAA80-肌动蛋白-脯氨酸蛋白三元复合物的晶体结构，代表不同的肌动蛋白同工型和催化反应的不同状态，并揭示了原子分辨率的NAT-底物复合物的第一结构。突变体的结构，生化和细胞分析表明，NAA80如何进化为可特异性识别所有细胞蛋白中的肌动蛋白，同时靶向所有六个肌动蛋白同工型，在氨基末端差异最大。