About PhosphoNET ...

PhosphoNET is an open-access, online resource developed by Kinexus Bioinformatics Corporation to foster the study of cell signalling systems to advance biomedical research in academia and industry. PhosphoNET is the worlds largest repository of known and predicted information on human phosphorylation sites, their evolutionary conservation, the identities of protein kinases that may target these sites and related phosphosites. PhosphoNET presently holds data on over 950,000 known and putative phosphorylation sites (P-sites) in over 20,000 human proteins that have been collected from the scientific literature and other reputable websites. Over 19% of these phosphosites have been experimentally validated. The rest have been predicted with a novel P-Site Predictor algorithm developed at Kinexus with academic partners at the University of British Columbia and Simon Fraser University.

With the PhosphoNET Evolution module, this website also provides information about cognate proteins in over 20 other species that may share these human phosphosites. This helps to define the most functionally important phosphosites as these are expected to be highly conserved in nature.

With the Kinase Predictor module, listings are provided for the top 50 human protein kinases that are likely to phosphorylate each of these phosphosites using another proprietary kinase substrate prediction algorithm developed at Kinexus.  Our kinase substrate predictions are based on deduced consensus phosphorylation site amino acid frequency scoring matrices that we have determined for each of ~500 different human protein kinases. The specificity matrices are generated directly from the primary amino acid sequences of the catalytic domains of these kinases, and when available, have proven to correlate strongly with substrate prediction matrices based on alignment of known substrates of these kinases. The higher the score, the better the prospect that a kinase will phosphorylate a given site. Over 30 million kinase-substrate phosphosite pairs are quantified in PhosphoNET. Kinexus Bioinformatics Corporation has the capability to test most of these putative interactions in vitro for our clients.

With the Phosphosite Match module, we have identified those phosphosites that show the greatest amino acid identity to a selected target human phosphosite. This tool is useful to identify related phosphosites that might be targeted the same protein kinases, phosphatases and/or adapter proteins. This type of query might also help identify potential off-targets for cross-reactive proteins that are detected with phosphosite-specific antibodies that were generated to recognize target phosphosites.


Role of Phosphorylation

Protein phosphorylation is well recognized as the premier mode of post-translational regulation of proteins. The human genome encodes over 20,000 proteins, and more than two thirds of these proteins have already been shown to be subject to phosphorylation. Research at Kinexus has revealed the vast majority of proteins are in fact phosphorylatable, and there may be very few exceptions. Based on the number of experimentally confirmed tyrosine phosphorylated sites in PhosphoNET (i.e. >36,000) and the percentage of tyrosine phosphorylation sites relative to the total number of phosphosites discovered in randomized studies (i.e. ~4%), we originally estimated that there are at least 500,000 human phosphosites. With our proprietary P-site Predictor algorithm, we have been able to identify over 800,000 new putative human phosphosites. Of the ~180,500 phosphosites that have been demonstrated, functional information is available for only about 1%. Only about 0.85% of these phosphorylations have been correlated with an increase in enzymatic activity or a gain of function, whereas only 0.3% result in a decrease in enzymatic activity or a reduction of function. We believe that only a small percentage of the human phosphorylation sites directly regulate activities of proteins and control protein-protein interactions. It would appear that the major role of hyperphosphorylation of proteins is actually to facilitate their degradation by proteases. The challenge for the biomedical research community is to identify the most critical phosphorylation sites that regulate protein functions and serve as biomarkers of human disease. Kinexus is committed to the identification of these important phosphosites and the development of tools to permit their analysis in experimental model and clinical specimens. PhosphoNET is a free Internet resource that has been developed to facilitate this important endeavor.



PhosphoNET is designed to be fast and simple to navigate. You can search for target protein if you know its UniProt ID, International Protein ID, or one of its common alias names. You can also search PhosphoNET with the complete 15 amino acid sequence (in single letter code) surrounding a phosphosite (the phosphorylated amino acid should be defined in the 8th position). A list of possible options for proteins are generated by typing at least three letters of their name or identification number, and waiting for a few seconds for a complete list to appear. Once a unique phosphoprotein webpage is created, you can hover your cursor over the coloured buttons to the right of the phosphosite sequences to get more information that will appear next to the button. If the button is clicked, this information will appear in the white Info Box. Clicking on the orange buttons activates links to additional webpages. PhosphoNET Evolution, PhosphoNET Kinase Predictor, and PhosphoNET Phosphosite Match are accessed by clicking on the orange buttons under the Evol., Kinase Pred., and "P-Site Match" headings, respectively. Dark grey buttons are presently inactive. The optimum web browsers for using PhosphoNET are Internet Explorer, Firefox and Safari.


Funding and Advertising Opportunities

To develop and sustain this free resource, we are seeking sponsorship from foundations and corporations. We gratefully acknowledge financial grant support for PhosphoNET from the National Research Council of Canada Industrial Assistance Program and the Natural Sciences and Engineering Research Council of Canada.

It is possible for commercial vendors of signal transduction reagents and services to advertise their relevant products and services on this site. Interested parties should contact our Sales and Marketing Department at or call toll free in North America 1-866-KINEXUS.


Some PhosphoNET Website Statistics

Total number of phosphosites: 966,817

Total number of experimentally-confirmed phosphosites: 177,424

Percent serine phosphosites: 56.0%

Percent threonine phosphosites: 23.5%

Percent tyrosine phosphosites: 20.4%

Total number of proteins represented: 22,698

Average number phosphosites per protein: 42.6

Number of phosphosites with some functional information: 1553

Number of phosphosites that are activatory: 631

Number of phosphosites that are inhibitory: 229

Average Mr of phosphoproteins: 61,520 Da

Median Mr of phosphoproteins: 46,000 Da

Average number amino acids per phosphoprotein: 552

Median number amino acids per phosphoprotein: 410


Copyright © 2019 Kinexus Bioinformatics Corporation