title: A crosslinker based on a tethered electrophile for mapping kinase-substrate networks.
abstract: Despite the continuing progress made toward mapping kinase signaling networks, there are still many phosphorylation events for which the responsible kinase has not yet been identified. We are interested in addressing this problem through forming covalent crosslinks between a peptide substrate and the corresponding phosphorylating kinase. Previously we reported a dialdehyde-based kinase-binding probe capable of such a reaction with a peptide containing a cysteine substituted for the phosphorylatable ser/thr/tyr residue. Here, we examine the yield of a previously reported dialdehyde-based probe and report that the dialdehyde-based probes possess a significant limitation in terms of crosslinked kinase-substrate product yield. To address this limitation, we developed a crosslinking scheme based on a kinase activity-based probe, and this crosslinker provides an increase in efficiency and substrate specificity, including in the context of cell lysate.
authors.affiliation: Howard Hughes Medical Institute, Department of Cellular and Molecular Pharmacology, University of California, San Francisco, 600 16(th) Street, MC2280, San Francisco, CA 94158, USA; Chemistry and Chemical Biology Graduate Program, University of California, San Francisco, 600 16(th) Street, MC2280, San Francisco, CA 94158, USA.
authors.forename: Megan M
authors.initials: MM
authors.lastname: Riel-Mehan
authors.affiliation: Chemistry and Chemical Biology Graduate Program, University of California, San Francisco, 600 16(th) Street, MC2280, San Francisco, CA 94158, USA. Electronic address: kevan.shokat@ucsf.edu.
authors.forename: Kevan M
authors.initials: KM
authors.lastname: Shokat
mesh_terms: D000447:Aldehydes / Q000737:chemistry / Q000378:metabolism*; D003432:Cross-Linking Reagents / Q000737:chemistry*; D015335:Molecular Probes / Q000737:chemistry / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D010770:Phosphotransferases / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D000447:Aldehydes; D003432:Cross-Linking Reagents; D015335:Molecular Probes; D010455:Peptides; D010770:Phosphotransferases
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