Functional Insights Into Protein Acetylation in the Hyperthermophilic Archaeon Sulfolobus islandicus*

Abstract

Proteins undergo acetylation at the Nε-amino group of lysine residues and the Nα-amino group of the N terminus in Archaea as in Bacteria and Eukarya. However, the extent, pattern and roles of the modifications in Archaea remain poorly understood. Here we report the proteomic analyses of a wild-type <i>Sulfolobus islandicus</i> strain and its mutant derivative strains lacking either a homolog of the protein acetyltransferase Pat (Δ<i>Sis</i>Pat) or a homolog of the Nt-acetyltransferase Ard1 (Δ<i>Sis</i>Ard1). A total of 1708 Nε-acetylated lysine residues in 684 proteins (26% of the total proteins), and 158 Nt-acetylated proteins (44% of the identified proteins) were found in <i>S. islandicus</i> Δ<i>Sis</i>Ard1 grew more slowly than the parental strain, whereas Δ<i>Sis</i>Pat showed no significant growth defects. Only 24 out of the 1503 quantifiable Nε-acetylated lysine residues were differentially acetylated, and all but one of the 24 residues were less acetylated by >1.3 fold in Δ<i>Sis</i>Pat than in the parental strain, indicating the narrow substrate specificity of the enzyme. Six acyl-CoA synthetases were the preferred substrates of <i>Sis</i>Pat <i>in vivo</i>, suggesting that Nε-acetylation by the acetyltransferase is involved in maintaining metabolic balance in the cell. Acetylation of acyl-CoA synthetases by <i>Sis</i>Pat occurred at a sequence motif conserved among all three domains of life. On the other hand, 92% of the acetylated N termini identified were acetylated by <i>Sis</i>Ard1 in the cell. The enzyme exhibited broad substrate specificity and could modify nearly all types of the target N termini of human NatA-NatF. The deletion of the <i>Sis</i>Ard1 gene altered the cellular levels of 18% of the quantifiable proteins (1518) by >1.5 fold. Consistent with the growth phenotype of Δ<i>Sis</i>Ard1, the cellular levels of proteins involved in cell division and cell cycle control, DNA replication, and purine synthesis were significantly lowered in the mutant than those in the parental strain.