3SVV: Crystal Structure Of T338c C-Src Covalently Bound To Vinylsulfonamide- Pyrazolopyrimidine 9

The conserved nature of the ATP-binding site of the > 500 human kinases renders the development of specific inhibitors a challenging task. A widely used chemical genetic strategy to overcome the specificity challenge exploits a large-to-small mutation of the gatekeeper residue (a conserved hydrophobic amino acid) and the use of a bulky inhibitor to achieve specificity via shape complementarity. However, in a number of cases, introduction of a glycine or alanine gatekeeper results in diminished kinase activity and ATP affinity. A new chemical genetic approach based on covalent complementarity between an engineered gatekeeper cysteine and an electrophilic inhibitor was developed to address these challenges. This strategy was evaluated with Src, a proto-oncogenic tyrosine kinase known to lose some enzymatic activity using the shape complementarity chemical genetic strategy. We found that Src with a cysteine gatekeeper recapitulates wild type activity and can be irreversibly inhibited both in vitro and in cells. A cocrystal structure of T338C c-Src with a vinylsulfonamide-derivatized pyrazolopyrimidine inhibitor was solved to elucidate the inhibitor binding mode. A panel of electrophilic inhibitors was analyzed against 307 kinases and MOK (MAPK/MAK/MRK overlapping kinase), one of only two human kinases known to have an endogenous cysteine gatekeeper. This analysis revealed remarkably few off-targets, making these compounds the most selective chemical genetic inhibitors reported to date. Protein engineering studies demonstrated that it is possible to increase inhibitor potency through secondary-site mutations. These results suggest that chemical genetic strategies based on covalent complementarity should be widely applicable to the study of protein kinases.
PDB ID: 3SVVDownload
MMDB ID: 92780
PDB Deposition Date: 2011/7/12
Updated in MMDB: 2011/10
Experimental Method:
x-ray diffraction
Resolution: 2.2  Å
Source Organism:
Similar Structures:
Biological Unit for 3SVV: monomeric; determined by author and by software (PISA)
Molecular Components in 3SVV
Label Count Molecule
Protein (1 molecule)
Proto-oncogene Tyrosine-protein Kinase SRC(Gene symbol: SRC)
Molecule annotation
Chemical (1 molecule)
* Click molecule labels to explore molecular sequence information.

Citing MMDB