Dataset Information

Chemical proteomics profiling of N-terminal and lysine acetyltransferases

ABSTRACT: Acetylation of amino groups is a prevalent protein modification in all kingdoms of life. Acetyl groups are transferred from Coenzyme A (CoA) to protein N-termini and lysine side chains by N-terminal acetyltransferases (NATs) and lysine acetyltransferases (KATs), respectively. Building on lysine-CoA conjugates as KAT probes, we have synthesized N-terminal CoA-conjugated peptide probes for interactome profiling of NAT complexes. These probes specifically recruited catalytic and auxiliary subunits of the major NAT complexes from cell lysates. When comparing the specificity of N-terminal and lysine CoA-conjugated probes the latter bound only a subset of endogenous KATs and appeared more prone to recruitment of other CoA-binding proteins. Western blot validation confirmed the specificity of selected NATs and KATs towards these probes supporting the notion that N-terminal CoA-conjugated peptides are versatile probes for NAT complex profiling in lysates of physiological and pathological backgrounds.

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Iris Finkemeier

PROVIDER: PXD018544 | JPOST Repository | Wed Apr 14 00:00:00 BST 2021

REPOSITORIES: jPOST

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Dataset's files

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			Action	DRS
	170214_C17_008_QHF_FH_Rev04_%2317_007_SER11_1.raw	Raw
	170214_C17_008_QHF_FH_Rev04_%2317_007_SER11_2.raw	Raw
	170214_C17_008_QHF_FH_Rev04_%2317_007_SER11_3.raw	Raw
	170214_C17_008_QHF_FH_Rev04_%2317_007_SER12_1.raw	Raw
	170214_C17_008_QHF_FH_Rev04_%2317_007_SER12_2.raw	Raw

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Publications

Investigating Peptide-Coenzyme A Conjugates as Chemical Probes for Proteomic Profiling of N-Terminal and Lysine Acetyltransferases.

Sindlinger Julia J Schön Stefan S Eirich Jürgen J Kirchgäßner Sören S Finkemeier Iris I Schwarzer Dirk D

Chembiochem : a European journal of chemical biology 20220725 17

Acetyl groups are transferred from acetyl-coenzyme A (Ac-CoA) to protein N-termini and lysine side chains by N-terminal acetyltransferases (NATs) and lysine acetyltransferases (KATs), respectively. Building on lysine-CoA conjugates as KAT probes, we have synthesized peptide probes with CoA conjugated to N-terminal alanine (α-Ala-CoA), proline (α-Pro-CoA) or tri-glutamic acid (α-3Glu-CoA) units for interactome profiling of NAT complexes. The α-Ala-CoA probe enriched the majority of NAT catalytic ...[more]

PMID: 35776679

Similar Datasets

Project description:Co-translational N-terminal (Nt-) acetylation of nascent polypeptides is catalyzed by N-terminal acetyltransferases (NATs). The very N-terminal amino acid sequence is the major factor determining whether or not a given protein is Nt-acetylated. In humans, six different NATs, denoted NatA-NatF, are identified. In the current study we used N-terminal COFRADIC analysis to define the in vivo substrate specificity of Naa50 (Nat5)/NatE as this has long remained elusive. Three yeast strains were generated; a control strain endogenously expressing yNaa50, a deletion strain depleted of yNaa50 and a strain deleted of yNaa50 but ectopically expressing human Naa50. When comparing the Nt-acetylation status of different N-termini in the control strain with the deletion strain, a reduction in Nt-acetylation for several yeast proteins was observed. To our surprise, these substrates were not of the predicted NatE-type substrates, but rather canonical NatA substrates. Further, ectopic expression of hNaa50 mainly resulted in the Nt-acetylation of a selected class of otherwise Nt-free yeast N-termini besides increasing the degree of Nt-acetylation of several other yeast proteins, and as such (partially) complementing those N-termini displaying reduced Nt-acetylation upon yNAA50-deletion. The preferred substrates of hNaa50 were predominantly Met-starting N-termini including Met-Lys-, Met-Val-, Met-Ala-, Met-Tyr-, Met-Phe-, Met-Leu-, Met-Ser- and Met-Thr, and highly overlapped with the previously identified human Naa60/NatF substrate specificity profile. Identification of several hNaa50 substrates with a small amino acid in the second position also revealed a potential interplay between the NATs and methionine aminopeptidases (MetAPs). The initiator Methionine (iMet) is normally cleaved off by MetAPs when the second amino acid is small, but our in vitro data suggest that in contrast to a free iMet, an Nt-acetylated iMet is not hydrolyzed by MetAPs. Thus, Naa50-mediated Nt-acetylation may potentially act as a mechanism to retain the iMet of proteins with a small amino acid at the second position that normally would be hydrolyzed.