CBI学会 2021年 大会, 口頭発表, O1-2, 21.10.26

1. Dynamic residue interaction network
analysis of the neuraminidase H274Y
mutant conferring drug resistance in
influenza virus.
Mohini YADAV (1) | Manabu IGARASHI (2) | Norifumi YAMAMOTO(1)
(1) Chiba Institute of Technology
(2) Hokkaido University
26th October 2021
O1-2 CBI Annual Meeting 2021

2. Zanamivir
Relenza
Oseltamivir
Tamiflu
Anti-Influenza Drugs
2

3. (Relenza) (Tamiflu)
Neuraminidase inhibitor mechanism
3

4. Detection of drug-resistant viruses in 2019/2020 influenza season (Japan)
Oseltamivir
(Tamiflu)
Zanamivir
(Relenza)
Resistant (%) 1.7 % 0 %
4
Source: National Institute of Infectious Diseases (NIID), Japan (August 18, 2021)
Anti-Influenza Drug Resistance

5. 5
Introduction: H274Y mutant of H5N1 avian influenza A virus
Source: Yen et al., Journal of virology, 2007; A/Vietnam/1203/04 (H5N1)
Mutation
(N2 numbering)
Oseltamivir
IC50 (nM)
Fold Change
WT 0.3 -
H274Y 501.7 1672

6. 6
Source: Collins et al., Nature, 2008
Introduction: H274Y mutant of H5N1 avian influenza A virus
Yellow – Wild type
Green – H274Y mutant

7. 7
Source: Malaisree et al., Amino Acids, 2009
Introduction: H274Y mutant of H5N1 avian influenza A virus

8. 8
AIM
AIM
To provide novel insights into the molecular mechanism underlying
oseltamivir drug resistance in influenza virus caused by
neuraminidase H274Y mutation using molecular dynamics
simulations.
PROBLEM
Previous studies focused only on clarifying the effect of H274Y mutation
on the interaction between oseltamivir and active site residues of
neuraminidase. However, the effect of H274Y mutation on the correlation
between neuraminidase active site and mutation site is still unclear.
In this study, the effect of H274Y mutation on the correlation
between neuraminidase active site and mutation site is clarified
using newly developed “Dynamic Residue Interaction Network
(dRIN) analysis”.
SOLUTION

9. • Hydrogen bond
• Van der Waals interactions
• Disulfide bridges
9
Source: Piovesan et al., Nucleic Acids Research, 2016
Residue Interaction Network (RIN)
• Salt bridges
• π-π stacking interactions
• π-cation interactions

10. Residue Interaction Network (RIN)
Hydrogen bond Van der Waals interactions Disulfide bridges
Salt bridges π-π stacking interactions π-cation interactions
10

11. Dynamic Residue Interaction Network (dRIN)
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X-Ray or NMR or MD simulation
Single protein structure
Residue Interaction Network
(RIN) analysis
Molecular Dynamics (MD) Simulation
Multiple protein structures
Dynamic Residue Interaction Network
(dRIN) analysis

12. Initial Structure PDB ID: 2HU4 (WT), 3CL0 (H274Y)
Force Field
Protein: ff14SB
Ligand (OTV): GAFF + RESP charge
Water: TIP3P
Condition NPT ensemble
Length 80 ns
Thermostat Weak-coupling algorithm
Barostat Berendsen algorithm
Software AMBER 20
Binding free energy
calculation
Molecular Mechanics Poisson Boltzmann Surface Area
(MM-PBSA)
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Method: Molecular Dynamics (MD) simulation

13. Dynamic Residue Interaction Network (dRIN)
13

14. Dynamic Residue Interaction Network (dRIN)
14

15. ∆G: -11.54 kcal mol−1 ∆G: -4.34 kcal mol−1
∆∆G: +7.20 kcal mol−1
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Result: Snapshot images and binding free energies

16. Result: Dynamic residue interaction network (dRIN) graphs
Hydrogen bond
Van der Waals interaction
π-π stacking interaction
Close contact
|||||||
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17. 17
Result: Superimposed snapshot images of the 150-loop region

18. • The H5N1 H274Y mutant is highly resistant to OTV.
• OTV binding site of NA and its H274Y mutation site interacts via three interface
residues, S246, E276 and R292.
• Due to H274Y mutation, interaction between residue 274 and interface residues
significantly increased, leading to significantly decreased interaction between OTV
and 150-loop residues, D151 and R152.
• This research work is published in PeerJ Journal.
18
Conclusion