Publication Abstract from PubMed
The upstream protein kinases responsible for thousands of phosphorylation events in the phosphoproteome remain to be discovered. We developed a three-component chemical reaction which converts the transient noncovalent substrate-kinase complex into a covalently cross-linked product by utilizing a dialdehyde-based cross-linker, 1. Unfortunately, the reaction of 1 with a lysine in the kinase active site and an engineered cysteine on the substrate to form an isoindole cross-linked product could not be performed in the presence of competing cellular proteins due to nonspecific side reactions. In order to more selectively target the cross-linker to protein kinases in cell lysates, we replaced the weak, kinase-binding adenosine moiety of 1 with a potent protein kinase inhibitor scaffold. In addition, we replaced the o-phthaldialdehyde moiety in 1 with a less-reactive thiophene-2,3-dicarboxaldehyde moiety. The combination of these two structural modifications provides for cross-linking of a cysteine-containing substrate to its corresponding kinase in the presence of competing cellular proteins.
Tuning a three-component reaction for trapping kinase substrate complexes.,Statsuk AV, Maly DJ, Seeliger MA, Fabian MA, Biggs WH 3rd, Lockhart DJ, Zarrinkar PP, Kuriyan J, Shokat KM J Am Chem Soc. 2008 Dec 24;130(51):17568-74. PMID:19053485[1]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
