PDBsum entry 1gag

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protein ligands metals Protein-protein interface(s) links
Transferase/transferase inhibitor PDB id
Protein chains
303 a.a. *
13 a.a. *
_MG ×2
Waters ×23
* Residue conservation analysis
PDB id:
Name: Transferase/transferase inhibitor
Title: Crystal structure of the insulin receptor kinase in complex bisubstrate inhibitor
Structure: Insulin receptor, tyrosine kinase domain. Chain: a. Fragment: tyrosine kinase domain. Synonym: ir. Engineered: yes. Mutation: yes. Bisubstrate peptide inhibitor. Chain: b. Engineered: yes.
Source: Homo sapiens. Human. Organism_taxid: 9606. Expressed in: spodoptera frugiperda. Expression_system_taxid: 7108. Synthetic: yes
Biol. unit: Dimer (from PQS)
2.70Å     R-factor:   0.214     R-free:   0.269
Authors: K.Parang,J.H.Till,A.J.Ablooglu,R.A.Kohanski,S.R.Hubbard,P.A.
Key ref:
K.Parang et al. (2001). Mechanism-based design of a protein kinase inhibitor. Nat Struct Biol, 8, 37-41. PubMed id: 11135668 DOI: 10.1038/83028
29-Nov-00     Release date:   17-Jan-01    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P06213  (INSR_HUMAN) -  Insulin receptor
1382 a.a.
303 a.a.*
Protein chain
No UniProt id for this chain
Struc: 13 a.a.
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: Chain A: E.C.  - Receptor protein-tyrosine kinase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: ATP + a [protein]-L-tyrosine = ADP + a [protein]-L-tyrosine phosphate
Bound ligand (Het Group name = 112)
matches with 83.33% similarity
+ [protein]-L-tyrosine
+ [protein]-L-tyrosine phosphate
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     membrane   1 term 
  Biological process     transmembrane receptor protein tyrosine kinase signaling pathway   2 terms 
  Biochemical function     transferase activity, transferring phosphorus-containing groups     5 terms  


DOI no: 10.1038/83028 Nat Struct Biol 8:37-41 (2001)
PubMed id: 11135668  
Mechanism-based design of a protein kinase inhibitor.
K.Parang, J.H.Till, A.J.Ablooglu, R.A.Kohanski, S.R.Hubbard, P.A.Cole.
Protein kinase inhibitors have applications as anticancer therapeutic agents and biological tools in cell signaling. Based on a phosphoryl transfer mechanism involving a dissociative transition state, a potent and selective bisubstrate inhibitor for the insulin receptor tyrosine kinase was synthesized by linking ATPgammaS to a peptide substrate analog via a two-carbon spacer. The compound was a high affinity competitive inhibitor against both nucleotide and peptide substrates and showed a slow off-rate. A crystal structure of this inhibitor bound to the tyrosine kinase domain of the insulin receptor confirmed the key design features inspired by a dissociative transition state, and revealed that the linker takes part in the octahedral coordination of an active site Mg2+. These studies suggest a general strategy for the development of selective protein kinase inhibitors.
  Selected figure(s)  
Figure 2.
Figure 2. Kinetic analysis of the inhibition of IRK by bisubstrate analog compound 2. a, E/V versus 1/ATP in the presence of varying concentrations of compound 2. b, E/V versus 1/IRS727 in the presence of varying compound 2. For (a), the apparent K[i] (compound 2) was 550 80 nM; for (b), the apparent K[i] (compound 2) was 750 90 nM. c, Product/Enzyme versus Time. This experiment monitors the phosphopeptide (product) formation after a rapid dilution of the enzyme -inhibitor complex to indirectly measure the k[off] for dissociation of compound 2 from IRK; k[off] = 0.013 0.001 s-1.
Figure 3.
Figure 3. Crystal structure of the binary complex between cIRK and the bisubstrate inhibitor. a, Overall view of the binary complex in which cIRK is shown in surface representation and the bisubstrate inhibitor in stick representation. The ATP S moiety of the inhibitor is colored green, the peptide moiety is colored red, and the linker connecting the nucleotide and peptide is colored yellow. The cIRK surface is semi-transparent; the N-terminal lobe of cIRK partially masks the nucleotide. b, Stereo view of the F[o] - F[c] electron density (2.7 resolution, 3 contour) for the bisubstrate inhibitor computed after simulated annealing (1,000 K), omitting from the atomic model either ATP S + linker (blue map) or the peptide moiety (green map). Coloring of the bisubstrate inhibitor is the same as in (a). Selected peptide residues are labeled; Y'(P0) refers to the modified Tyr at the P0 position of the peptide. The purple sphere represents the Mg2+ and the red sphere represents the Mg2+-coordinating water molecule. c, Stereo view of the interactions between the inhibitor and key catalytic residues. Superimposed are the cIRK -bisubstrate inhibitor (binary) complex and the cIRK -MgAMP-PNP -peptide (ternary) complex15. Oxygen atoms are colored red, nitrogen atoms blue, sulfur atoms green, and phosphorus atoms yellow. Bonds/carbon atoms are colored orange for the binary complex and green for the ternary complex. Bonds and atoms of the ternary complex are semi-transparent. Mg2+ ions and water molecules are represented as purple and red spheres, respectively. Hydrogen bonds and bonds to the Mg2+ are represented as dashed and solid black lines, respectively. Only the modified tyrosine from the peptide moiety of the inhibitor is shown. 'BSI' indicates the bisubstrate inhibitor in the binary complex, and 'Y(P0)' and 'PNP' indicate the substrate tyrosine and AMP-PNP, respectively, in the ternary complex. Selected secondary structural elements ( C and 3) are shown. The figure was prepared with GRASP28, BOBSCRIPT29 and MOLSCRIPT30.
  The above figures are reprinted by permission from Macmillan Publishers Ltd: Nat Struct Biol (2001, 8, 37-41) copyright 2001.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21097901 B.P.Barnett, Y.Hwang, M.S.Taylor, H.Kirchner, P.T.Pfluger, V.Bernard, Y.Y.Lin, E.M.Bowers, C.Mukherjee, W.J.Song, P.A.Longo, D.J.Leahy, M.A.Hussain, M.H.Tschöp, J.D.Boeke, and P.A.Cole (2010).
Glucose and weight control in mice with a designed ghrelin O-acyltransferase inhibitor.
  Science, 330, 1689-1692.  
19774589 D.Lavogina, E.Enkvist, and A.Uri (2010).
Bisubstrate inhibitors of protein kinases: from principle to practical applications.
  ChemMedChem, 5, 23-34.  
20237675 J.van Ameijde, A.J.Poot, L.T.van Wandelen, A.E.Wammes, R.Ruijtenbeek, D.T.Rijkers, and R.M.Liskamp (2010).
Preparation of novel alkylated arginine derivatives suitable for click-cycloaddition chemistry and their incorporation into pseudosubstrate- and bisubstrate-based kinase inhibitors.
  Org Biomol Chem, 8, 1629-1639.  
20520657 R.Huang, I.Martinez-Ferrando, and P.A.Cole (2010).
Enhanced interrogation: emerging strategies for cell signaling inhibition.
  Nat Struct Mol Biol, 17, 646-649.  
19618415 A.J.Poot, J.van Ameijde, M.Slijper, A.van den Berg, R.Hilhorst, R.Ruijtenbeek, D.T.Rijkers, and R.M.Liskamp (2009).
Development of selective bisubstrate-based inhibitors against protein kinase C (PKC) isozymes by using dynamic peptide microarrays.
  Chembiochem, 10, 2042-2051.  
19674907 C.D.Shomin, S.C.Meyer, and I.Ghosh (2009).
Staurosporine tethered peptide ligands that target cAMP-dependent protein kinase (PKA): optimization and selectivity profiling.
  Bioorg Med Chem, 17, 6196-6202.  
19433564 D.H.Fong, and A.M.Berghuis (2009).
Structural basis of APH(3')-IIIa-mediated resistance to N1-substituted aminoglycoside antibiotics.
  Antimicrob Agents Chemother, 53, 3049-3055.
PDB codes: 3h8p 3tm0
19260709 K.E.Muratore, M.A.Seeliger, Z.Wang, D.Fomina, J.Neiswinger, J.J.Havranek, D.Baker, J.Kuriyan, and P.A.Cole (2009).
Comparative analysis of mutant tyrosine kinase chemical rescue.
  Biochemistry, 48, 3378-3386.
PDB code: 3geq
19632467 L.M.Szewczuk, M.K.Tarrant, and P.A.Cole (2009).
Protein phosphorylation by semisynthesis: from paper to practice.
  Methods Enzymol, 462, 1.  
19489734 M.K.Tarrant, and P.A.Cole (2009).
The chemical biology of protein phosphorylation.
  Annu Rev Biochem, 78, 797-825.  
19731277 R.Tiwari, and K.Parang (2009).
Protein conjugates of SH3-domain ligands and ATP-competitive inhibitors as bivalent inhibitors of protein kinases.
  Chembiochem, 10, 2445-2448.  
18282484 B.E.Turk (2008).
Understanding and exploiting substrate recognition by protein kinases.
  Curr Opin Chem Biol, 12, 4.  
18396877 K.A.Pickin, S.Chaudhury, B.C.Dancy, J.J.Gray, and P.A.Cole (2008).
Analysis of protein kinase autophosphorylation using expressed protein ligation and computational modeling.
  J Am Chem Soc, 130, 5667-5669.  
18800048 P.A.Cole (2008).
Chemical probes for histone-modifying enzymes.
  Nat Chem Biol, 4, 590-597.  
17530729 A.Kumar, Y.Wang, X.Lin, G.Sun, and K.Parang (2007).
Synthesis and Evaluation of 3-Phenylpyrazolo[3,4-d]pyrimidine-Peptide Conjugates as Src Kinase Inhibitors.
  ChemMedChem, 2, 1346-1360.  
18000050 K.Arora, and C.L.Brooks (2007).
Large-scale allosteric conformational transitions of adenylate kinase appear to involve a population-shift mechanism.
  Proc Natl Acad Sci U S A, 104, 18496-18501.  
17522047 S.Y.Ku, P.Yip, K.A.Cornell, M.K.Riscoe, J.B.Behr, G.Guillerm, and P.L.Howell (2007).
Structures of 5-methylthioribose kinase reveal substrate specificity and unusual mode of nucleotide binding.
  J Biol Chem, 282, 22195-22206.
PDB codes: 2pu8 2pui 2pul 2pun 2pup
17674392 Y.Ahmadibeni, M.Hanley, M.White, M.Ayrapetov, X.Lin, G.Sun, and K.Parang (2007).
Metal-binding properties of a dicysteine-containing motif in protein tyrosine kinases.
  Chembiochem, 8, 1592-1605.  
16843266 D.K.Walters, T.Mercher, T.L.Gu, T.O'Hare, J.W.Tyner, M.Loriaux, V.L.Goss, K.A.Lee, C.A.Eide, M.J.Wong, E.P.Stoffregen, L.McGreevey, J.Nardone, S.A.Moore, J.Crispino, T.J.Boggon, M.C.Heinrich, M.W.Deininger, R.D.Polakiewicz, D.G.Gilliland, and B.J.Druker (2006).
Activating alleles of JAK3 in acute megakaryoblastic leukemia.
  Cancer Cell, 10, 65-75.  
16669643 J.M.Hah, V.Sharma, H.Li, and D.S.Lawrence (2006).
Acquisition of a "Group A"-selective Src kinase inhibitor via a global targeting strategy.
  J Am Chem Soc, 128, 5996-5997.  
16640460 N.M.Levinson, O.Kuchment, K.Shen, M.A.Young, M.Koldobskiy, M.Karplus, P.A.Cole, and J.Kuriyan (2006).
A Src-like inactive conformation in the abl tyrosine kinase domain.
  PLoS Biol, 4, e144.
PDB codes: 2g1t 2g2f 2g2h 2g2i
17085043 R.Bose, M.A.Holbert, K.A.Pickin, and P.A.Cole (2006).
Protein tyrosine kinase-substrate interactions.
  Curr Opin Struct Biol, 16, 668-675.  
16777603 X.Zhang, J.Gureasko, K.Shen, P.A.Cole, and J.Kuriyan (2006).
An allosteric mechanism for activation of the kinase domain of epidermal growth factor receptor.
  Cell, 125, 1137-1149.
PDB codes: 2gs2 2gs6 2gs7
16226484 D.Schwarzer, and P.A.Cole (2005).
Protein semisynthesis and expressed protein ligation: chasing a protein's tail.
  Curr Opin Chem Biol, 9, 561-569.  
15003245 C.Luo, and P.Laaja (2004).
Inhibitors of JAKs/STATs and the kinases: a possible new cluster of drugs.
  Drug Discov Today, 9, 268-275.  
14978303 M.R.Ferguson, X.Fan, M.Mukherjee, J.Luo, R.Khan, J.C.Ferreon, V.J.Hilser, R.E.Shope, and R.O.Fox (2004).
Directed discovery of bivalent peptide ligands to an SH3 domain.
  Protein Sci, 13, 626-632.  
14570903 M.Y.Niv, H.Rubin, J.Cohen, L.Tsirulnikov, T.Licht, A.Peretzman-Shemer, E.Cna'an, A.Tartakovsky, I.Stein, S.Albeck, I.Weinstein, M.Goldenberg-Furmanov, D.Tobi, E.Cohen, M.Laster, S.A.Ben-Sasson, and H.Reuveni (2004).
Sequence-based design of kinase inhibitors applicable for therapeutics and target identification.
  J Biol Chem, 279, 1242-1255.  
15551269 P.De Meyts (2004).
Insulin and its receptor: structure, function and evolution.
  Bioessays, 26, 1351-1362.  
12573349 C.B.Gambacorti-Passerini, R.H.Gunby, R.Piazza, A.Galietta, R.Rostagno, and L.Scapozza (2003).
Molecular mechanisms of resistance to imatinib in Philadelphia-chromosome-positive leukaemias.
  Lancet Oncol, 4, 75-85.  
12547913 E.Chapman, M.C.Bryan, and C.H.Wong (2003).
Mechanistic studies of beta-arylsulfotransferase IV.
  Proc Natl Acad Sci U S A, 100, 910-915.  
12954329 G.Scapin, S.B.Patel, J.Lisnock, J.W.Becker, and P.V.LoGrasso (2003).
The structure of JNK3 in complex with small molecule inhibitors: structural basis for potency and selectivity.
  Chem Biol, 10, 705-712.
PDB codes: 1pmn 1pmq 1pmu 1pmv
14690593 J.Hu, J.Liu, R.Ghirlando, A.R.Saltiel, and S.R.Hubbard (2003).
Structural basis for recruitment of the adaptor protein APS to the activated insulin receptor.
  Mol Cell, 12, 1379-1389.
PDB codes: 1rpy 1rqq
12044161 A.Cook, E.D.Lowe, E.D.Chrysina, V.T.Skamnaki, N.G.Oikonomakos, and L.N.Johnson (2002).
Structural studies on phospho-CDK2/cyclin A bound to nitrate, a transition state analogue: implications for the protein kinase mechanism.
  Biochemistry, 41, 7301-7311.
PDB code: 1gy3
12047871 G.Scapin (2002).
Structural biology in drug design: selective protein kinase inhibitors.
  Drug Discov Today, 7, 601-611.  
12191607 K.Parang, and P.A.Cole (2002).
Designing bisubstrate analog inhibitors for protein kinases.
  Pharmacol Ther, 93, 145-157.  
12043997 S.E.Martin, and B.R.Peterson (2002).
A colorimetric enzyme-linked on-bead assay for identification of synthetic substrates of protein tyrosine kinases.
  J Pept Sci, 8, 227-233.  
12138114 S.Munshi, M.Kornienko, D.L.Hall, J.C.Reid, L.Waxman, S.M.Stirdivant, P.L.Darke, and L.C.Kuo (2002).
Crystal structure of the Apo, unactivated insulin-like growth factor-1 receptor kinase. Implication for inhibitor specificity.
  J Biol Chem, 277, 38797-38802.
PDB code: 1m7n
11598120 A.J.Ablooglu, M.Frankel, E.Rusinova, J.B.Ross, and R.A.Kohanski (2001).
Multiple activation loop conformations and their regulatory properties in the insulin receptor's kinase domain.
  J Biol Chem, 276, 46933-46940.  
11694888 S.Favelyukis, J.H.Till, S.R.Hubbard, and W.T.Miller (2001).
Structure and autoregulation of the insulin-like growth factor 1 receptor kinase.
  Nat Struct Biol, 8, 1058-1063.
PDB code: 1k3a
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