Protein sequencing by mass analysis of polypeptide ladders after controlled protein hydrolysis Export

@article{citeulike:5788837, abstract = {The characterization of protein modifications is essential for the study of protein function using functional genomic and proteomic approaches. However, current techniques are not efficient in determining protein modifications. We report an approach for sequencing proteins and determining modifications with high speed, sensitivity and specificity. We discovered that a protein could be readily acid-hydrolyzed within 1 min by exposure to microwave irradiation to form, predominantly, two series of polypeptide ladders containing either the N- or C-terminal amino acid of the protein, respectively. Mass spectrometric analysis of the hydrolysate produced a simple mass spectrum consisting of peaks exclusively from these polypeptide ladders, allowing direct reading of amino acid sequence and modifications of the protein. As examples, we applied this technique to determine protein phosphorylation sites as well as the sequences and several previously unknown modifications of 28 small proteins isolated from Escherichia coli K12 cells. This technique can potentially be automated for large-scale protein annotation.}, author = {Zhong, Hongying and Zhang, Ying and Wen, Zhihui and Li, Liang}, citeulike-article-id = {5788837}, citeulike-linkout-0 = {http://dx.doi.org/10.1038/nbt1011}, citeulike-linkout-1 = {http://dx.doi.org/10.1038/nbt1011}, day = {07}, doi = {10.1038/nbt1011}, journal = {Nature Biotechnology}, keywords = {n-terminal-peptides, proteomics, tandem-mass-spectrometry}, month = {September}, number = {10}, pages = {1291--1296}, posted-at = {2009-09-15 18:02:06}, priority = {2}, publisher = {Nature Publishing Group}, title = {Protein sequencing by mass analysis of polypeptide ladders after controlled protein hydrolysis}, url = {http://dx.doi.org/10.1038/nbt1011}, volume = {22}, year = {2004} }

TY - JOUR ID - citeulike:5788837 L3 - citeulike-article-id:5788837 N2 - The characterization of protein modifications is essential for the study of protein function using functional genomic and proteomic approaches. However, current techniques are not efficient in determining protein modifications. We report an approach for sequencing proteins and determining modifications with high speed, sensitivity and specificity. We discovered that a protein could be readily acid-hydrolyzed within 1 min by exposure to microwave irradiation to form, predominantly, two series of polypeptide ladders containing either the N- or C-terminal amino acid of the protein, respectively. Mass spectrometric analysis of the hydrolysate produced a simple mass spectrum consisting of peaks exclusively from these polypeptide ladders, allowing direct reading of amino acid sequence and modifications of the protein. As examples, we applied this technique to determine protein phosphorylation sites as well as the sequences and several previously unknown modifications of 28 small proteins isolated from Escherichia coli K12 cells. This technique can potentially be automated for large-scale protein annotation. IS - 10 JF - Nature Biotechnology EP - 1296 TI - Protein sequencing by mass analysis of polypeptide ladders after controlled protein hydrolysis PB - Nature Publishing Group VL - 22 SP - 1291 KW - n-terminal-peptides KW - proteomics KW - tandem-mass-spectrometry AU - Zhong, Hongying AU - Zhang, Ying AU - Wen, Zhihui AU - Li, Liang PY - 2004/09/07/ UR - http://dx.doi.org/10.1038/nbt1011 ER -