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International audience; N-(α)-terminal protein acetylation (NTA) is a widespread and essential protein modification. The reaction is catalyzed by various N-acetyltransferases (Nats) composed of a catalytic subunit alone or in complex with one or several factors. Their substrate specificities are largely determined by the first two N-terminal amino acids of the substrate protein. NTA is considered as an irreversible mark and mostly occurs co-translationally. A multiplicity of NTA roles have been uncovered such as its involvement in influencing protein's half-life. However, the majority of the proteome undergoes NTA with a high stoichiometry suggesting that NTA fulfils a broader and collective function, yet to be discovered. In this context, it has been suggested that NTA participates in maintaining protein homeostasis or proteostasis. The nucleus is rich in metastable and stress sensitive proteins suggesting that specific protein quality control strategies are in place to maintain nuclear proteostasis. Indeed

Large-scale biology article; International audience; N-terminal myristoylation, a major eukaryotic protein lipid modification, is difficult to detect in vivo and challenging to predict in silico. We developed a proteomics strategy involving subfractionation of cellular membranes, combined with separation of hydrophobic peptides by mass spectrometry-coupled liquid chromatography to identify the Arabidopsis thaliana myristoylated proteome. This approach identified a starting pool of 8837 proteins in all analyzed cellular fractions, comprising 32% of the Arabidopsis proteome. Of these, 906 proteins contain an N-terminal Gly at position 2, a prerequisite for myristoylation, and 214 belong to the predicted myristoylome (comprising 51% of the predicted myristoylome of 421 proteins). We further show direct evidence of myristoylation in 72 proteins; 18 of these myristoylated proteins were not previously predicted. We found one myristoylation site downstream of a predicted initiation codon, indicating that posttranslational myristoylation occurs in plants. Over half of the identified proteins could be quantified and assigned to a subcellular compartment. Hierarchical clustering of protein accumulation combined with myristoylation and S-acylation data revealed that N-terminal double acylation influences redirection to the plasma membrane. In a few cases, MYR function extended beyond simple membrane association. This study identified hundreds of N-acylated proteins for which lipid modifications could control protein localization and expand protein function.

International audience; Prokaryotic proteins must be deformylated before the removal of their first methionine. Peptide deformylase (PDF) is indispensable and guarantees this mechanism. Recent metagenomics studies revealed new idiosyncratic PDF forms as the most abundant family of viral sequences. Little is known regarding these viral PDFs, including the capacity of the corresponding encoded proteins to ensure deformylase activity. We provide here the first evidence that viral PDFs, including the shortest PDF identified to date, Vp16 PDF, display deformylase activity in vivo, despite the absence of the key ribosome-interacting C-terminal region. Moreover, characterization of phage Vp16 PDF underscores unexpected structural and molecular features with the C-terminal Isoleucine residue significantly contributing to deformylase activity both in vitro and in vivo. This residue fully compensates for the absence of the usual long C-domain. Taken together, these data elucidate an unexpected mechanism of enzyme natural evolution and adaptation within viral sequences.

EnCOUNTer parameters file. (PDF 1Â kb)

Distribution of the transit peptide cleavage position for fraction 5 validated dataset (True and False) and few specific subset (Mitochondria, plastidâ Ś). (PDF 120Â kb)

Web logos and heatmaps of 231 of nuclear coded plastidic N-termini (A-B), 30 mitochondrial N-termini (C-D) and 35Â N-termini associated to protein carrying an excised signal peptide (E-F). The sequences around the transit peptide cleavage are compared to the average distribution of A. thaliana using the stand alone ICELogo tool [41]. (PDF 474Â kb)

Mascot Distiller processing method for protein N-terminal acetylation quantitation. (PDF 24Â kb)

tMCC profiles of the Spec matrix for each possible residue for the A) True subset, B) False subset and C) from random A. thaliana proteins. (PDF 382Â kb)