PDBsum entry 1kmf

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Hormone/growth factor PDB id
Protein chains
21 a.a.
30 a.a. *
* Residue conservation analysis
PDB id:
Name: Hormone/growth factor
Title: Nmr structure of human insulin mutant ile-a2-allo-ile, his- b10-asp, pro-b28-lys, lys-b29-pro, 15 structures
Structure: Insulin. Chain: a. Engineered: yes. Mutation: yes. Insulin. Chain: b. Engineered: yes. Mutation: yes
Source: Synthetic: yes. Other_details: the peptide was chemically synthesized. The sequence of the peptide is naturally found in homo sapiens (human).. (Human).
NMR struc: 15 models
Authors: B.Xu,Q.X.Hua,S.H.Nakagawa,W.Jia,Y.C.Chu,P.G.Katsoyannis, M.A.Weiss
Key ref:
B.Xu et al. (2002). Chiral mutagenesis of insulin's hidden receptor-binding surface: structure of an allo-isoleucine(A2) analogue. J Mol Biol, 316, 435-441. PubMed id: 11866509 DOI: 10.1006/jmbi.2001.5377
14-Dec-01     Release date:   09-Jan-02    
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Protein chain
Pfam   ArchSchema ?
P01308  (INS_HUMAN) -  Insulin
110 a.a.
21 a.a.*
Protein chain
Pfam   ArchSchema ?
P01308  (INS_HUMAN) -  Insulin
110 a.a.
30 a.a.*
Key:    PfamA domain  Secondary structure
* PDB and UniProt seqs differ at 4 residue positions (black crosses)


DOI no: 10.1006/jmbi.2001.5377 J Mol Biol 316:435-441 (2002)
PubMed id: 11866509  
Chiral mutagenesis of insulin's hidden receptor-binding surface: structure of an allo-isoleucine(A2) analogue.
B.Xu, Q.X.Hua, S.H.Nakagawa, W.Jia, Y.C.Chu, P.G.Katsoyannis, M.A.Weiss.
The hydrophobic core of vertebrate insulins contains an invariant isoleucine residue at position A2. Lack of variation may reflect this side-chain's dual contribution to structure and function: Ile(A2) is proposed both to stabilize the A1-A8 alpha-helix and to contribute to a "hidden" functional surface exposed on receptor binding. Substitution of Ile(A2) by alanine results in segmental unfolding of the A1-A8 alpha-helix, lower thermodynamic stability and impaired receptor binding. Such a spectrum of perturbations, although of biophysical interest, confounds interpretation of structure-activity relationships. To investigate the specific contribution of Ile(A2) to insulin's functional surface, we have employed non-standard mutagenesis: inversion of side-chain chirality in engineered monomer allo-Ile(A2)-DKP-insulin. Although the analogue retains native structure and stability, its affinity for the insulin receptor is impaired by 50-fold. Thus, whereas insulin's core readily accommodates allo-isoleucine at A2, its activity is exquisitely sensitive to chiral inversion. We propose that the Ile(A2) side-chain inserts within a chiral pocket of the receptor as part of insulin's hidden functional surface.
  Selected figure(s)  
Figure 1.
Figure 1. Insulin's ``hidden'' functional-surface hypothesis and corresponding CD studies. (a) Sche- matic model proposing confor- mational change on receptor binding, exposing Ile A2 and novel non-polar surface. (b) Cylinder model of insulin T-state. 5,6 Detach- ment of C-terminal B-chain b- strand on receptor binding would expose ``hidden'' surface spanning Ile A2 and Val A3 . 2,8,23 Native or ``closed'' form of B chain is anchored by engagement of Phe B24 against hydrophobic core. (c) Far- UV CD spectra of DKP-insulin (continuous line), allo-Ile A2 -DKP- insulin (*), and Ala A2 -DKP-insulin (*) suggest attenuated helix con- tent in Ala A2 -DKP-insulin 25 but native helix content in allo-Ile A2 - DKP-insulin. (d) Thermal unfold- ing, monitored by ellipticity at 222 nm, demonstrates native stab- ility of allo-Ile A2 -DKP-insulin (*) relative to DKP-insulin (continuous line) whereas Ala A2 -DKP-insulin (*) is less stable. 25 (e) Guanidine unfolding transitions of Ala A2 -DKP- insulin (*), allo-Ile A2 -DKP-insulin (*) and DKP-insulin (continuous line) demonstrate that whereas Ala A2 -DKP variant exhibits decreased stability (deltadeltaGu 0.4-1.2 kcal/mol), 25 allo-Ile A2 -DKP-insulin has similar or greater stability (see footnote { on page 437). Two-state modeling and extracted thermodynamic parameters are provided in Supplementary Material. CD samples contained 25-50 mM insulin or analogue in 50 mM potassium phosphate (pH 7); samples were diluted to 5 mM for equilibrium denaturation studies.
Figure 4.
Figure 4. Core packing schemes. (a) Environment of Ile A2 (black) in T-state crystal structures in relation to Val A3 , Leu A16 , Tyr A19 , Leu B11 , Leu B15 , and Tyr B26 . (b) and (c) Corresponding interactions of A2 side-chain in NMR-derived ensembles of (b) DKP-insulin ensemble and (c) allo-Ile A2 -DKP-insulin. Except for chiral inversion of its b-carbon, packing interactions of allo-Ile A2 are simi- lar to those of Ile A2 in crystal and solution structures. Structures in each panel are aligned with respect to main-chain atoms of A2-A8, A13-A20 and B9-B19. Excepting A2, A-chain side-chains are shown in red and B-chain side-chains in blue.
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2002, 316, 435-441) copyright 2002.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20133841 J.Jirácek, L.Záková, E.Antolíková, C.J.Watson, J.P.Turkenburg, G.G.Dodson, and A.M.Brzozowski (2010).
Implications for the active form of human insulin based on the structural convergence of highly active hormone analogues.
  Proc Natl Acad Sci U S A, 107, 1966-1970.
PDB codes: 2wru 2wrv 2wrw 2wrx 2ws0 2ws1 2ws4 2ws6 2ws7
20948967 M.Liu, L.Haataja, J.Wright, N.P.Wickramasinghe, Q.X.Hua, N.F.Phillips, F.Barbetti, M.A.Weiss, and P.Arvan (2010).
Mutant INS-gene induced diabetes of youth: proinsulin cysteine residues impose dominant-negative inhibition on wild-type proinsulin transport.
  PLoS One, 5, e13333.  
20106974 Y.Yang, Q.X.Hua, J.Liu, E.H.Shimizu, M.H.Choquette, R.B.Mackin, and M.A.Weiss (2010).
Solution structure of proinsulin: connecting domain flexibility and prohormone processing.
  J Biol Chem, 285, 7847-7851.
PDB code: 2kqp
19321435 B.Xu, K.Huang, Y.C.Chu, S.Q.Hu, S.Nakagawa, S.Wang, R.Y.Wang, J.Whittaker, P.G.Katsoyannis, and M.A.Weiss (2009).
Decoding the Cryptic Active Conformation of a Protein by Synthetic Photoscanning: INSULIN INSERTS A DETACHABLE ARM BETWEEN RECEPTOR DOMAINS.
  J Biol Chem, 284, 14597-14608.  
19321436 Q.X.Hua, B.Xu, K.Huang, S.Q.Hu, S.Nakagawa, W.Jia, S.Wang, J.Whittaker, P.G.Katsoyannis, and M.A.Weiss (2009).
Enhancing the Activity of a Protein by Stereospecific Unfolding: CONFORMATIONAL LIFE CYCLE OF INSULIN AND ITS EVOLUTIONARY ORIGINS.
  J Biol Chem, 284, 14586-14596.
PDB codes: 2k91 2k9r
18332129 Q.X.Hua, S.H.Nakagawa, W.Jia, K.Huang, N.B.Phillips, S.Q.Hu, and M.A.Weiss (2008).
Design of an active ultrastable single-chain insulin analog: synthesis, structure, and therapeutic implications.
  J Biol Chem, 283, 14703-14716.
PDB codes: 2jzq 3bxq
18492668 Z.L.Wan, K.Huang, S.Q.Hu, J.Whittaker, and M.A.Weiss (2008).
The structure of a mutant insulin uncouples receptor binding from protein allostery. An electrostatic block to the TR transition.
  J Biol Chem, 283, 21198-21210.  
17410596 J.P.Mayer, F.Zhang, and R.D.DiMarchi (2007).
Insulin structure and function.
  Biopolymers, 88, 687-713.  
17884811 K.Huang, S.J.Chan, Q.X.Hua, Y.C.Chu, R.Y.Wang, B.Klaproth, W.Jia, J.Whittaker, P.De Meyts, S.H.Nakagawa, D.F.Steiner, P.G.Katsoyannis, and M.A.Weiss (2007).
The A-chain of insulin contacts the insert domain of the insulin receptor. Photo-cross-linking and mutagenesis of a diabetes-related crevice.
  J Biol Chem, 282, 35337-35349.
PDB codes: 2jum 2juu 2juv
17716170 M.Koch, F.F.Schmid, V.Zoete, and M.Meuwly (2006).
Insulin: a model system for nanomedicine?
  Nanomed, 1, 373-378.  
16894147 M.Lou, T.P.Garrett, N.M.McKern, P.A.Hoyne, V.C.Epa, J.D.Bentley, G.O.Lovrecz, L.J.Cosgrove, M.J.Frenkel, and C.W.Ward (2006).
The first three domains of the insulin receptor differ structurally from the insulin-like growth factor 1 receptor in the regions governing ligand specificity.
  Proc Natl Acad Sci U S A, 103, 12429-12434.
PDB code: 2hr7
16728398 Q.X.Hua, M.Liu, S.Q.Hu, W.Jia, P.Arvan, and M.A.Weiss (2006).
A conserved histidine in insulin is required for the foldability of human proinsulin: structure and function of an ALAB5 analog.
  J Biol Chem, 281, 24889-24899.
PDB code: 2h67
16762918 Q.X.Hua, S.Nakagawa, S.Q.Hu, W.Jia, S.Wang, and M.A.Weiss (2006).
Toward the active conformation of insulin: stereospecific modulation of a structural switch in the B chain.
  J Biol Chem, 281, 24900-24909.
PDB codes: 2hh4 2hho
16751187 S.H.Nakagawa, Q.X.Hua, S.Q.Hu, W.Jia, S.Wang, P.G.Katsoyannis, and M.A.Weiss (2006).
Chiral mutagenesis of insulin. Contribution of the B20-B23 beta-turn to activity and stability.
  J Biol Chem, 281, 22386-22396.  
16080143 V.Zoete, M.Meuwly, and M.Karplus (2005).
Study of the insulin dimerization: binding free energy calculations and per-residue free energy decomposition.
  Proteins, 61, 79-93.  
14988398 Q.X.Hua, and M.A.Weiss (2004).
Mechanism of insulin fibrillation: the structure of insulin under amyloidogenic conditions resembles a protein-folding intermediate.
  J Biol Chem, 279, 21449-21460.
PDB code: 1sf1
14596591 Z.L.Wan, B.Xu, Y.C.Chu, P.G.Katsoyannis, and M.A.Weiss (2003).
Crystal structure of allo-Ile(A2)-insulin, an inactive chiral analogue: implications for the mechanism of receptor binding.
  Biochemistry, 42, 12770-12783.
PDB codes: 1lw8 1pc1 1q4v
12196530 Q.X.Hua, Y.C.Chu, W.Jia, N.F.Phillips, R.Y.Wang, P.G.Katsoyannis, and M.A.Weiss (2002).
Mechanism of insulin chain combination. Asymmetric roles of A-chain alpha-helices in disulfide pairing.
  J Biol Chem, 277, 43443-43453.
PDB code: 1lkq
12198117 U.Narendra, L.Zhu, B.Li, J.Wilken, and M.A.Weiss (2002).
Sex-specific gene regulation. The Doublesex DM motif is a bipartite DNA-binding domain.
  J Biol Chem, 277, 43463-43473.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB codes are shown on the right.