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BibTex format

author = {Goya, Grocin A and Serwa, R and Morales, Sanfrutos J and Ritzefeld, M and Tate, E},
doi = {10.1074/mcp.RA118.001043},
journal = {Molecular and Cellular Proteomics},
pages = {115--126},
title = {Whole proteome profiling of N-myristoyltransferase activity and inhibition using Sortase A},
url = {},
volume = {18},
year = {2019}

RIS format (EndNote, RefMan)

AB - N-myristoylation is the covalent addition of a 14-carbon saturated fatty acid (myristate) to the N-terminal glycine of specific protein substrates by N-myristoyltransferase (NMT) and plays an important role in protein regulation by controlling localization, stability, and interactions. We developed a novel method for whole-proteome profiling of free N-terminal glycines through labeling with S. Aureus sortase A (SrtA) and used it for assessment of target engagement by an NMT inhibitor. Analysis of the SrtA-labeling pattern with an engineered biotinylated depsipeptide SrtA substrate (Biotin-ALPET-Haa, Haa = 2-hydroxyacetamide) enabled whole proteome identification and quantification of de novo generated N-terminal Gly proteins in response to NMT inhibition by nanoLC-MS/MS proteomics, and was confirmed for specific substrates across multiple cell lines by gel-based analyses and ELISA. To achieve optimal signal over background noise we introduce a novel and generally applicable improvement to the biotin/avidin affinity enrichment step by chemically dimethylating commercial NeutrAvidin resin and combining this with two-step LysC on-bead/trypsin off-bead digestion, effectively eliminating avidin-derived tryptic peptides and enhancing identification of enriched peptides. We also report SrtA substrate specificity in whole-cell lysates for the first time, confirming SrtA promiscuity beyond its recognized preference for N-terminal glycine, and its usefulness as a tool for unbiased labeling of N-terminal glycine-containing proteins. Our new methodology is complementary to metabolic tagging strategies, providing the first approach for whole proteome gain-of signal readout for NMT inhibition in complex samples which are not amenable to metabolic tagging.
AU - Goya,Grocin A
AU - Serwa,R
AU - Morales,Sanfrutos J
AU - Ritzefeld,M
AU - Tate,E
DO - 10.1074/mcp.RA118.001043
EP - 126
PY - 2019///
SN - 1535-9476
SP - 115
TI - Whole proteome profiling of N-myristoyltransferase activity and inhibition using Sortase A
T2 - Molecular and Cellular Proteomics
UR -
UR -
VL - 18
ER -