An N α-Acetyltransferase Responsible for Acetylation of the N-terminal Residues of Histones H4 and H2A

Abstract

A yeast gene has been identified that encodes a novel, evolutionarily conserved N ␣ -acetyltransferase responsible for acetylation of the N-terminal residues of histones H4 and H2A. The gene has been named NAT4. Recombinant Nat4 protein acetylated a peptide corresponding to the N-terminal tail of H4, but not an H3 peptide nor the peptide adrenocorticotropin. H4 and H2A are N-terminally acetylated in all species from yeast to mammals and hence blocked from sequencing by Edman degradation. In contrast, H4 and H2A purified from a nat4 mutant were unacetylated and could be sequenced. Analysis of yeast histones by acid-urea gel electrophoresis showed that all the H4 and H2A from the mutant migrated more rapidly than the same histones from a wild type strain, consistent with the histones from the mutant having one extra positive charge due to one less acetylated amino group. A comparison of yeast proteins from wild type and a nat4 mutant by two-dimensional gel electrophoresis showed no evidence that other yeast proteins are substrates of this acetyltransferase. Thus, Nat4 may be dedicated specifically to the N-terminal acetylation of histones H4 and H2A. Surprisingly, nat4 mutants grow at a normal rate and have no readily observable phenotypes.Eukaryotic proteins are subject to two cotranslational modifications as the nascent polypeptides emerge from the ribosome, methionine cleavage by methionine aminopeptidase, and acetylation of the ␣-amino group of the N-terminal amino acid (1). Removal of the methionine occurs only if the second amino acid is a small one such as serine, alanine, or glycine. Acetylation of the ␣-amino group on either methionine, or on the N-terminal residue resulting from methionine cleavage, occurs in the great majority of (but not all) eukaryotic proteins, and is accomplished by one of several N ␣ -acetyltransferases (NATs) 1 (1). A number of years ago we identified the genes for a yeast NAT, now called NatA, that consists of two subunits, Ard1 and Nat1 (2). Subsequent work showed that NatA was responsible for the acetylation of many, but not all, yeast proteins beginning with small residues such as serine, alanine, or glycine (2-5). Yeast ard1 and nat1 mutants are viable and the many proteins that are no longer N-terminally acetylated in the mutant strains are as stable as they are in a wild type strain (2). Thus, there is no evidence that N-terminal acetylation serves to protect proteins from degradation in yeast.Ard1 protein has an acetyl-CoA binding motif found in members of the GNAT superfamily, enzymes that acetylate amino groups on proteins and other molecules (6). Thus, Ard1 is very likely to be the catalytic subunit of NatA. Two other NATs have been identified in yeast, called NatB and NatC, with catalytic subunits Nat3 and Mak3, respectively (4, 7). NatB and NatC both acetylate proteins with N-terminal methionine residues, although they have different specificities dictated by the nature of the subsequent amino acids (4).Histone H4 is a highly conserved protein with an N-terminal serin...

Antimicrobial Components of the Neonatal Gut Affected Upon Colonization

“…Attempts to determine the N-terminal sequence of these polypeptides failed, suggesting that they were N-terminally blocked. Hence, they were identified with peptide mass fingerprinting, and found to be histone H4 and H2A, which are indeed N-terminally blocked (20). Histone H4 was identified with eight matching fragments in 62% total sequence coverage, and histone H2A with seven matching fragments in 40% total sequence coverage.…”

Combinatorial Modification of Human Histone H4 Quantitated by Two-dimensional Liquid Chromatography Coupled with Top Down Mass Spectrometry

“…Unacetylated, Methylated H4-It is well known that all H4 molecules are co-translationally acetylated on the ␣-NH 2 of Ser-1 (56,57), resulting in a ⌬m of at least ϩ42 Da for all forms of H4. This ϩ42 Da form will be referred to as the "unacetylated" form of H4 (a␣Ser-1), whereas the term "monoacety-FIGURE 1.…”

Histone H4 Acetylation Differentially Modulates Arginine Methylation by an in Cis Mechanism

“…To quantitatively evaluate the in cis effect of acetylation marks on Arg-3 methylation, we designed a library of H4 peptides containing the first 20 amino acids that incorporate all 16 possible acetylation combinations, including 1 unacetylated, 4 monoacetylated, 6 diacetylated, 4 triacetylated, and 1 tetraacetylated forms (Table 1). All peptides are N-terminally capped with acetic anhydride because virtually all H4 proteins are N-terminally acetylated in vivo (10,21,22,49). These peptides are used as substrates of PRMT1 and PRMT5 to evaluate how different acetylation combinations affect the methylation of Arg-3 catalyzed by these two enzymes.…”