Although experimental data provide useful information, it still remains difficult to identify the true N-terminal peptides amid the multitude of internal peptides identified in a large scale experiment. The cleavage positions of the mitochondrial or plastid TP (mTP or cTP) can be predicted using TargetP or ChloroP softwares, but the predictions are not always reliable. The determination of the associated cleavage site of the transit peptide (TP) are still challenging. Large scale analyses show that 20–40% of these proteins are N-acetylated in their mature chloroplastic form.
In plants, most chloroplast proteins are encoded in the nucleus, translated in the cytosol and targeted to the chloroplast by a transit peptide that is cleaved upon arrival inside the organelle. N-terminal acetylation (NTA) is one of the major protein modifications of the eukaryotic cytosol and occurs mainly co-translationally. In this context, we have developed the EnCOUNTer tool (i) to score all characterized peptides using discriminating parameters to identify bona fide mature protein N-termini and (ii) to determine the N-terminus acetylation yield of the most reliable ones.
To date, no bioinformatics tool is available to define mature protein starting positions, the associated N-terminus acetylation status and/or yield for each proteoform. Although N-terminal acetylation occurs mostly in the cytosol, it is also observed in some organelles such as chloroplast. The situation becomes even more complex when post-translational modifications take place at the neo-N-terminus.
#Termius error codes manual#
Although some bioinformatics tools are available, they usually require extensive manual validation to identify the exact N-terminal position. Protein neo-N-termini can be located up to 100–150 amino acids downstream from the initiator methionine and are not easily predictable. This is especially true for proteins undergoing cleavage of transit peptides when they are targeted to specific organelles, such as mitochondria or chloroplast. Characterization of mature protein N-termini by large scale proteomics is challenging.
0 kommentar(er)
