1. PTP catalytic mechanism
the general acid
the general base

2. WPD loop
D to A
mutant

3. DA (trapping)
PTPs
WT (active)
PTPs

4. Background PTP1B Knock Out Mice
PTP1B -/- mice display enhanced sensitivity to insulin
PTP1B -/- mice are resistant to development of obesity and Type 2 diabetes
(Elchebly et al. Science (1999) 83, ) (Klaman et al. Mol Cell Biol (2000) 20, )

5. Inhibition of the Insulin Receptor PTP may Enhance Insulin Signaling
Prolonged or
Enhanced Signal
Transient
Signal

6. Structure of the PTP1B-IR peptide complex
G1166
R1164
Phe182
Arg24
Arg254
pY1163
pY1162
Asp48
D1161
Tyr46
To understand What makes the bis and tris such good PTP1B substrate we solved the structure of PTP1B in complex with the IR peptides.
Now, this is a surface representation of PTP1B in complex with the bisphospho peptide.
The recognition of the substrate phosphotyrosine 1162 is similar to that for the EGFR-peptide where the pTYr extend into the catalytic site.
The binding of the substrate pTyr induces closure of the WPD loop with Phen182 and Tyr46 sandwiching the tyrosine ring of pTYr 1162.
As in the PTP1B-EGFR complex the depth of the catalytic cleft is defined by Tyr 46 and Nterminal acidic residues interact with PTP1B and orient the peptide
Thus, Asp 1161 of the IR peptide interacts with and Arg47.
The length of pTyr1163 to make the salt bridge interactions with Arg24 and 254
The amide groups of pTYr 1162 and 1163 interacts with the carboxylate side chain of Asp 48.
Now What makes the bis and tris phophopeptides such good substrates is the salt bridge interactions Arg24 and 254 with the phosphoryl group of 1163 – In constratrs to the active site phosphotyrosine there is little interactions the phenyl ring
From structural alignment studies PTP1B appears to be unique in having a wide cleft that allows access of peptides to this second binding site.
In PTPalpha and LAR – which also have been implicated in IR signalling – this cleft is blocked by bulky residues and these PTPs would not recognizes a tandem pTYr motif with high affinity.
Finally, at the very C-terminal Arg1164 forms a pi-cation interaction with the phenyl ring of Phe182 of the WPD loop which requires the loop to be closed ans Gly1166 is also defined in the structure.
Arg47

7. IRTK Activation segment:
DIpY1158ETDpY1162pY1163RKGG
S.R. Hubbard, (1997) EMBO J., 16,

8. Kinetic Constants for Dephosphorylation of
IR Activation Loop Peptide by PTP1B
IR-peptide Km
[mM]
pY-1158
> 100
pY-1162
pY-1163
pY-1162,1163
14 ± 1.1
pY-1158,
1162,1163
7.9 ± 1.3
RDIpY1158ETDpY1162pY1163RKGGKGLL

9. PTP1B and insulin signaling
Insulin Receptor
PTK
Signaling
Response
PTP1B

10. Suppression of PTP1B expression by RNAi
may enhance tyrosine phosphorylation of IR
972
1158
PTP
1162
1163
Prolonged or
Enhanced Signal
Transient
Signal