Delia Turner '08 and Blaise Milburn '07
Insulin kinase receptor (IRK) is a transmembrane receptor that
begins insulin's signaling cascade that controls growth and metabolism.
IRK is an α2β2 glycoprotein and a member of the receptor tyrosine
kinase family. Insulin binds to the extracellular α chains causing
structural changes and autophosphorylation of cytoplasmic tyrosine residues
of the β chain. This phosphorylation causes recruitment of insulin
receptor signaling (IRS) proteins, which further propagate the insulin signaling
pathway (Li et al. 2005).
Protein tyrosine phosphatases (PTPs) dephosphorylate receptor tyrosine
kinases, thereby maintaining basal-level phosphorylation in cells. Protein
tyrosine phosphatase 1B (PTP1B) is one of the few PTPs known to dephosphorylate
IRK. The cornerstone of many cell signaling events rests on reversible phosphorylation
of tyrosine residues on proteins, including IRK in the insulin signaling
pathway (Stoker 2005). PTP1B is widely expressed and localized in the endoplasmic
reticulum. It has a clearable carboxy terminal that allows release into
the cytosol. From there, PTP1B binds to the backside of the kinase domain
rather to a phosphotyrosine in the activation loop "front side".
In this way, PTP1B acts as a negative regulator of insulin action, preventing
cells from becoming overly sensitive to insulin.
II. Overall Structure
The PTP1B-IRK complex is an asymmetric dimer consisting of 2 PTP1B-IRK
dimers interacting through an IRK-IRK interface.
Each PTP1B molecule is composed of 9 α-helices
and 9 β-strands.
Two very important loops within the PTP1B molecules are the PTP or catalytic
loops and the WPD loops.
Each IRK molecule is composed 9 α-helices
and 12 β-strands.
Two important components of the IRK molecules are the catalytic
loop and the activation loop
which contains the phosphorylated Tyr residues (Tyr 1158, 1162, and 1163)
. It is interesting to note that the PTP1Bs do not interact with the activation
loop of the IRKs, but rather the backsides of the IRKs. This has important
implications for the role of PTP1B.
The interface is nearly identical on both copies of PTP1B-IRK. There are 2 main regions of interaction encompassing both N and C terminal lobes of IRK: β2-β3 loop of IRK with β9-β10 turn of PTP1B, and the α helix J of IRK with the WPD loop (between β11 and α2) of PTP1B
. The key interaction in the first region between PTP1B and IRK is the hydrogen bond between E1022 (β2-β3 loop of IRK)
and Y152 (β9-β10 turn of PTP1B)
. Additionally, N193 (α3 of PTP1B)
forms a hydrogen bond with the backbone of G1021 (IRK)
, and Y153 (β10 of PTP1B)
forms a van der Waals contact with the backbone of G1021 (IRK)
The WPD loop of PTP1B (residues 179-184) makes multiple contacts with the αJ helix of IRK. Important
hydrophobic packing interactions in this region include P180 (WPD loop)
with V1274 (αJ)
and F182 (WPD loop)
with P1269 (αJ)
. Other interactions include a salt bridge between K120 (β6-β7 loop of PTP1B)
and E1273 (αJ)
, a hydrogen bond between R112 (β4-β5 loop of PTP1B)
and the backbone of E1273 (αJ)
, and a hydrogen bond between H1268 (αI-αJ loop of IRK)
and the backbone of F182 (PTP1B)
. The interactions between αJ and the WPD loop promote a closed conformation
of this catalytically important loop, which remains in the open conformation
when a phosphotyrosine is not bound in the catalytic site.
The IRK molecules form an asymmetric dimmer interacting through the N-terminal domain. The helix αC is the site of the primary contacts; van der Waals contact occurs between 4 hydrophobic residues (L1038, I1042, L1045, and A1048) that interlock with the antiparallel corresponding residues of the other IRK molecule.
Several hydrogen bonds involving the aC helix also help to stabilize the dimer interface. R1041 of aC forms hydrogen bonds with the backbone ß4-ß5 loop of the opposing IRK molecule and also with the side chain of the ß4-ß5 loop Q1070 residue.
Additionally, a hydrogen bond is formed between the ß3-aC loop of one IRK molecule and the ß4-ß5 loop of the other
The crystal structure PTP1B-IRK molecules represents a noncatalytic mode of interaction. This is indicated by the backside, rather than active site, binding of PTP1B to IRK. The occurrence of a noncatalytic interaction is supported by a FRET experiment (Romsicki 2004). It was found that a PTP1b catalytic site inhibitor did not decrease the basal FRET signal of PTP1b/insulin receptor interaction. This suggests that PTP1B not only dephosphorylates the insulin receptor, but also helps to maintain the dephosphorylation in a noncatalytic setting.
It has been suggested that PTP1B also binds to the backside of
unphosphorylated insulin receptor (Li 2005). This could allow the
PTP1b to dephosphorylate the insulin receptor more quickly upon its phosphorylation.
This is supported by faster PTP1b’s faster association time with the
insulin receptor than with other growth factor receptors. An alternative
hypothesis for backside binding is a result of the IRK-IRK dimerization.
PTP1b has a high affinity for phosphorylated insulin receptor. Therefore,
PTP1b would not bind to the backside of the kinase domain unless it was
not able to bind to the active site and the phosphorylated tyrosines.
The dimerization of IRK-IRK protects the phosphotyrosines while PTP1b alternatively
binds to the backside of the kinase domain.
The backside binding of the insulin receptor kinase domain introduces a new avenue of interest in growth and cellular metabolism regulation signaling pathways. Further investigation should explore the function of backside binding and its place in the regulation of insulin regulation.
Hubbard, S.R. 1997. Crystal structure of the activated insulin
receptor tyrosine kinase in complex with peptide substrate and ATP analog.
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Li S., Depetris R.S., Barford D., Chernoff J., Hubbard S.R. 2005.
Crystal Structure of a Complex between Protein Tyrosine Phosphatase 1B and
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Romsicki Y., Reece M., Gauthier J.Y., Asante-Appiah E., Kennedy
B.P. 2004. Protein tyrosine phosphatase-1B dephosphorylation of the
insulin receptor occurs in a perinuclear endosome compartment in human embryonic
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Salmeen A., Andersen J.N., Myers M.P., Tonks N.K., Barford D. 2000.
Molecular basis for the dephosphorylation of the activation segment of the
insulin receptor by protein tyrosine phosphatase 1B. Mol. Cell
Stoker, A. 2005. Protein tyrosine phosphatases and signaling. Journal
of Endocrinology 185:19-33.
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