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DIABETES CAUSED BY MUTANT INSULINS (CHICAGO, LOS
ANGELES AND WAKAYAMA) IN HUMANS: HOW DO THEY
BIND TO THE INSULIN RECEPTOR?
Md Ataul Islam, Tahir S. Pillay
Email: [email protected], [email protected] (*Funded by the UP Vice Chancellor’s
Postdoctoral Fellowship)
Department of Chemical Pathology, Faculty of Health Science,
University of Pretoria and NHLS-Tshwane Academic Division,
Pretoria, South Africa
Objective
1) Understand the molecular basis of decreased
receptor binding of mutant insulins which cause
diabetes in humans
2) Use molecular docking and molecular dynamics
studies to design higher affinity insulin derivatives
with altered pharmacological/biological properties
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Introduction
The crystal structure of human insulin bound to the extracellular domain of
the insulin receptor was recently reported in 2014.
There are a number of naturally occurring mutations in the insulin molecule in
humans which are associated with diabetes mellitus due to defective receptor
binding.
Diabetes associated mutations have been identified at three invariant sites:
 insulin Wakayama (Val3A → Leu3A)
 insulin Los Angeles (Phe24B → Ser24B)
 insulin Chicago (Phe25B→Leu25B)
There are no crystal structures between mutant insulins and IR till date.
Protein-protein docking and molecular dynamics are well established tools to
explore protein-protein interactions that play a major role in cellular
processes.
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Insulin Wakayama
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Insulin Wakayama
Normal Insulin
Val3 replaced by Leu3
Val3A at natural insulin
Leu3A at insulin Wakayama
Insulin Los Angeles
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Normal Insulin
Insulin Los Angeles
Phe24 replaced by Ser24
Phe24B at natural insulin
Ser24B at insulin Los Angeles
Insulin Chicago
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Normal Insulin
Insulin Chicago
Phe25 replaced by Leu25
Phe25B at natural insulin
Leu25B at Chicago insulin
Insulin Receptor (IR)
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Domain
Amino residue range
L1
1 – 157
CR
158 – 310
L2
311 – 470
FnIII – 1
471 – 595
FnIII – 2
596 – 808
FnIII – 3
809 -906
αCT
704 - 719
Protein-protein docking
Protein-protein interactions play important roles in many essential
biological processes, such as signal transduction, transport, cell
regulation, enzyme inhibition, etc.
These interactions very often lead to the formation of stable
protein–protein complex that are essential to perform their
biological functions.
Protein-protein docking finds the computationally relative 8
transformation and energetically favourable stable complex.
ZDOCK server was used for protein-protein docking
Online tool Contact Map Analysis (CMA) was considered for
binding interaction analysis.
Work flow
Natural insulin downloaded from PDB
Mutation by DeepView tool:
 Val3A → Leu3A
 Phe24B → Ser24B
 Phe25B→Leu25B
Mutant insulins
1. insulin Wakayama
2. insulin Los Angeles
3. insulin Chicago
Molecular Docking using
ZDOCK server
Best docked pose
Analyze of binding interactions
Molecular Dynamics
Analysis of binding interaction
and stability of complex
ZDOCK server
ZDOCK is the Fast Fourier Transform based protein docking
program.
ZDOCK searches all possible binding modes in the translational
and rotational space between the two proteins and evaluates each
pose using an energy-based scoring function.
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ZDOCK server
home page
From the crystal structure of insulin bound IR it was observed that
• The side chain of His710 of αCT inserts into a pocket formed by insulin
residues Val3A, Gly8B, Ser9B, and Val12B.
• The Phe714 interacted with insulin residues Gly1A, Ile2A, Tyr19A, Leu11B,
Val12B, and Leu15B.
• The side chain of Asn711 is directed toward with insulin residues Gly1A,
Val3A, and Glu4A.
• The αCT helix appears to an an interaction with the insulin A-chain Cterminus.
• The side chain of insulin residue Val12B is positioned between L1 domain
residues Phe39, Phe64, and Arg65, while that of insulin residue Tyr16B 11
adjoins that of the L1 domain residue Phe39.
Binding interaction analysis
Insulin Wakayama
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Insulin Los Angeles
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There was no role of Phe24B of insulin molecule to form
connection with IR.
When Phe24B amino acid changed to Ser24B (Los Angeles
insulin), played an important role to establish the complex by
forming interactions with Phe39 and Lys40 through 13 and 8
connections respectively.
Insulin Chicago
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Phe25B was not able to form interactions with IR in crystal
structure (3W14)
After mutation by Leu25B (Chicago insulin) and docked to IR, it
was observed that Leu25B was successfully established
connections with Asn15 and Ans16 belongs to L1 domain of IR.
Molecular dynamics
• Molecular dynamics gives the precise binding interactions at the
active site.
• Desmond of Schrodinger was use for molecular dynamics study.
•
• The Root Mean Square Deviation (RMSD) is used to measure the
average change in displacement of a selection of atoms for a
particular frame with respect to a reference frame.
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• The Root Mean Square Fluctuation (RMSF) is useful for
characterizing local changes along the protein chain.
• B-factors are the simplest method to analyze local deformability.
RMSD vs. Time
Normal insulin
Los Angeles insulin
Wakayama insulin
Chicago insulin
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RMSF vs. Residue index
αCT
FnIII - 1
CR
Wakayama insulin
Normal insulin
CR
FnIII - 1
αCT
FnIII - 1
CR
αCT
Los Angeles insulin
L2, FnIII -1
αCT
Chicago insulin
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Conclusions
It can be concluded that owing to the change of side chains in
the mutant insulins a substantial decrement of binding
interactions is observed.
The molecular dynamics study also indicated the instability of
the docked complex of mutant insulins to crystal structure..
Observation of molecular docking and molecular dynamics may
guide to design some higher affinity insulin derivatives with
altered pharmacological properties.
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Acknowledgment
• Prof. Tahir S. Pillay
• University of Pretoria
Postdoctoral Fellowship
Vice
Chancellor’s
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References
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