Actin’s acetylated N terminus is prominently exposed in the actin filament where it can interact with many filamentous actin-binding proteins (ABPs) ( 9), and consistently, actin Nt-acetylation critically affects filament assembly in vitro and in cells. NAA80 is highly specific, targeting only actin among all the proteins in the cell, whereas its knockout (KO) completely abolishes actin Nt-acetylation, showing that no other NAT can substitute for NAA80. Then, all six actin isoforms, which at this point carry either three (cytoplasmic β/γ-actin and smooth muscle γ-actin) or four (muscle α-actin isoforms) negatively charged amino acids at the N terminus, are posttranslationally Nt-acetylated by NAA80 ( 7, 10). This is followed by the removal of the N-terminal Ac-Met (or Ac-Cys) by a still unidentified acetylaminopeptidase. During translation, the initiator methionine (iMet) of the two cytoplasmic actin isoforms is Nt-acetylated, whereas the iMet of the four-muscle actin isoforms is removed and the exposed cysteine is Nt-acetylated by one of the ribosome-associated NATs. Actin Nt-acetylation is also unique in that it proceeds through a multistep mechanism that involves both co- and posttranslational modification ( 8, 9). In addition, we recently found that actin, the most abundant protein in the cytoplasm of eukaryotic cells, has its own dedicated NAT, NAA80 ( 7). One exception is NatD, which Nt-acetylates only two substrates, histones H2A and H4 ( 6). Most NATs are rather promiscuous, acting on large subsets of proteins.
Five of the NATs (NatA to NatE) associate with the ribosome and act cotranslationally, whereas NatF binds to the Golgi membrane and Nt-acetylates transmembrane proteins ( 5).
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The NATs transfer the acetyl group (COCH 3) from acetyl coenzyme A (AcCoA) onto the free N-terminal α-amino group of their protein substrates.
The reaction is catalyzed by one of seven Nt-acetyltransferases (NATs) (NatA to NatF and NatH), whose catalytic subunits (NAA10 to NAA60 and NAA80) display a common fold but low sequence identity. N-terminal acetylation (Nt-acetylation) is a prevalent modification that affects ~80% of human proteins and can have a major impact on human health and disease ( 1– 4). 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. 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. NAA80 Nt-acetylates actin-profilin much more efficiently than actin alone, suggesting that profilin acts as a chaperone for actin Nt-acetylation. 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.
Actin, the most abundant protein in the cytoplasm, has its own dedicated NAT, NAA80, which acts posttranslationally and affects cytoskeleton assembly and cell motility. About 80% of human proteins are amino-terminally acetylated (Nt-acetylated) by one of seven Nt-acetyltransferases (NATs).