Discovery of a new phosphotyrosine mimetic for PTP1B using breakaway tethering.
Protein tyrosine phosphatases play important roles in many signaling cascades involved in human disease. The identification of druglike inhibitors for these targets is a major challenge, and the discovery of suitable phosphotyrosine (pY) mimetics remains one of the key difficulties. Here we describe an extension of tethering technology, "breakaway tethering", which is ideally suited for discovering such new chemical entities. The approach involves first irreversibly modifying a protein with an extender that contains both a masked thiol and a known pY mimetic. The extender is then cleaved to release the pY mimetic, unmasking the thiol. The resulting protein is screened against a library of disulfide-containing small molecule fragments; any molecules with inherent affinity for the pY binding site will preferentially form disulfides with the extender, allowing for their identification by mass spectrometry. The ability to start from a known substrate mimimizes perturbation of protein structure and increases the opportunity to probe the active site using tethering. We applied this approach to the anti-diabetic protein PTP1B to discover a pY mimetic which belongs to a new molecular class and which binds in a novel fashion.
Erlanson
Daniel A
DA
Sunesis Pharmaceuticals, Inc., 341 Oyster Point Boulevard, South San Francisco, California 94080, USA. erlanson@sunesis.com
McDowell
Robert S
RS
He
Molly M
MM
Randal
Mike
M
Simmons
Robert L
RL
Kung
Jenny
J
Waight
Andrew
A
Hansen
Stig K
SK
D001665:Binding Sites; D040761:Biomimetic Materials / Q000737:chemistry* / Q000378:metabolism; D018360:Crystallography, X-Ray; D003545:Cysteine / Q000737:chemistry; D008958:Models, Molecular; D019815:Oxalic Acid / Q000737:chemistry; D019000:Phosphotyrosine / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry* / Q000378:metabolism
D019000:Phosphotyrosine; D019815:Oxalic Acid; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases; D003545:Cysteine