Mutations in the fusion protein of measles virus that confer resistance to the membrane fusion inhibitors carbobenzoxy-D-Phe-L-Phe-Gly and 4-nitro-2- phenylacetyl amino-benzamide

Michael N. Ha, Sébastien Delpeut, Ryan S. Noyce, Gary Sisson, Karen M. Black, Liang Tzung Lin, Darius Bilimoria, Richard K. Plemper, Gilbert G. Privé, Christopher D. Richardson

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

The inhibitors carbobenzoxy (Z)-D-Phe-L-Phe-Gly (fusion inhibitor peptide [FIP]) and 4-nitro-2-phenylacetyl amino-benzamide (AS-48) have similar efficacies in blocking membrane fusion and syncytium formation mediated by measles virus (MeV). Other homologues, such as Z-D-Phe, are less effective but may act through the same mechanism. In an attempt to map the site of action of these inhibitors, we generated mutant viruses that were resistant to the inhibitory effects of Z-D-Phe-LPhe- Gly. These 10 mutations were localized to the heptad repeat B (HRB) region of the fusion protein, and no changes were observed in the viral hemagglutinin, which is the receptor attachment protein. Mutations were validated in a luciferase-based membrane fusion assay, using transfected fusion and hemagglutinin expression plasmids or with syncytium-based assays in Vero, Vero-SLAM, and Vero-Nectin 4 cell lines. The changes I452T, D458N, D458G/V459A, N462K, N462H, G464E, and I483R conferred resistance to both FIP and AS-48 without compromising membrane fusion. The inhibitors did not block hemagglutinin protein-mediated binding to the target cell. Edmonston vaccine/laboratory and IC323 wild-type strains were equally affected by the inhibitors. Escape mutations were mapped upon a three-dimensional (3D) structure modeled from the published crystal structure of parainfluenzavirus 5 fusion protein. The most effective mutations were situated in a region located near the base of the globular head and its junction with the alpha-helical stalk of the prefusion protein. We hypothesize that the fusion inhibitors could interfere with the structural changes that occur between the prefusion and postfusion conformations of the fusion protein.

Original languageEnglish
Article numbere01026-17
JournalJournal of Virology
Volume91
Issue number23
DOIs
Publication statusPublished - Dec 1 2017

Fingerprint

glycyl-glycyl-glycyl-glycine
benzamides
Measles virus
Membrane Fusion
mutation
Mutation
Hemagglutinins
hemagglutinins
Giant Cells
phenylalanylglycine
Proteins
Viral Hemagglutinins
proteins
giant cells
Peptides
Protein Conformation
Luciferases
Protein Binding
peptides
Plasmids

Keywords

  • Antiviral inhibitors
  • AS-48
  • Drug resistance
  • Escape mutations
  • F protein
  • FIP
  • Fusion inhibitors
  • Measles virus
  • Membrane fusion
  • Z-D-Phe-L-Phe-Gly

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

Cite this

Mutations in the fusion protein of measles virus that confer resistance to the membrane fusion inhibitors carbobenzoxy-D-Phe-L-Phe-Gly and 4-nitro-2- phenylacetyl amino-benzamide. / Ha, Michael N.; Delpeut, Sébastien; Noyce, Ryan S.; Sisson, Gary; Black, Karen M.; Lin, Liang Tzung; Bilimoria, Darius; Plemper, Richard K.; Privé, Gilbert G.; Richardson, Christopher D.

In: Journal of Virology, Vol. 91, No. 23, e01026-17, 01.12.2017.

Research output: Contribution to journalArticle

Ha, Michael N. ; Delpeut, Sébastien ; Noyce, Ryan S. ; Sisson, Gary ; Black, Karen M. ; Lin, Liang Tzung ; Bilimoria, Darius ; Plemper, Richard K. ; Privé, Gilbert G. ; Richardson, Christopher D. / Mutations in the fusion protein of measles virus that confer resistance to the membrane fusion inhibitors carbobenzoxy-D-Phe-L-Phe-Gly and 4-nitro-2- phenylacetyl amino-benzamide. In: Journal of Virology. 2017 ; Vol. 91, No. 23.
@article{b55507656c984fd2ac3887f8f223656b,
title = "Mutations in the fusion protein of measles virus that confer resistance to the membrane fusion inhibitors carbobenzoxy-D-Phe-L-Phe-Gly and 4-nitro-2- phenylacetyl amino-benzamide",
abstract = "The inhibitors carbobenzoxy (Z)-D-Phe-L-Phe-Gly (fusion inhibitor peptide [FIP]) and 4-nitro-2-phenylacetyl amino-benzamide (AS-48) have similar efficacies in blocking membrane fusion and syncytium formation mediated by measles virus (MeV). Other homologues, such as Z-D-Phe, are less effective but may act through the same mechanism. In an attempt to map the site of action of these inhibitors, we generated mutant viruses that were resistant to the inhibitory effects of Z-D-Phe-LPhe- Gly. These 10 mutations were localized to the heptad repeat B (HRB) region of the fusion protein, and no changes were observed in the viral hemagglutinin, which is the receptor attachment protein. Mutations were validated in a luciferase-based membrane fusion assay, using transfected fusion and hemagglutinin expression plasmids or with syncytium-based assays in Vero, Vero-SLAM, and Vero-Nectin 4 cell lines. The changes I452T, D458N, D458G/V459A, N462K, N462H, G464E, and I483R conferred resistance to both FIP and AS-48 without compromising membrane fusion. The inhibitors did not block hemagglutinin protein-mediated binding to the target cell. Edmonston vaccine/laboratory and IC323 wild-type strains were equally affected by the inhibitors. Escape mutations were mapped upon a three-dimensional (3D) structure modeled from the published crystal structure of parainfluenzavirus 5 fusion protein. The most effective mutations were situated in a region located near the base of the globular head and its junction with the alpha-helical stalk of the prefusion protein. We hypothesize that the fusion inhibitors could interfere with the structural changes that occur between the prefusion and postfusion conformations of the fusion protein.",
keywords = "Antiviral inhibitors, AS-48, Drug resistance, Escape mutations, F protein, FIP, Fusion inhibitors, Measles virus, Membrane fusion, Z-D-Phe-L-Phe-Gly",
author = "Ha, {Michael N.} and S{\'e}bastien Delpeut and Noyce, {Ryan S.} and Gary Sisson and Black, {Karen M.} and Lin, {Liang Tzung} and Darius Bilimoria and Plemper, {Richard K.} and Priv{\'e}, {Gilbert G.} and Richardson, {Christopher D.}",
year = "2017",
month = "12",
day = "1",
doi = "10.1128/JVI.01026-17",
language = "English",
volume = "91",
journal = "Journal of Virology",
issn = "0022-538X",
publisher = "American Society for Microbiology",
number = "23",

}

TY - JOUR

T1 - Mutations in the fusion protein of measles virus that confer resistance to the membrane fusion inhibitors carbobenzoxy-D-Phe-L-Phe-Gly and 4-nitro-2- phenylacetyl amino-benzamide

AU - Ha, Michael N.

AU - Delpeut, Sébastien

AU - Noyce, Ryan S.

AU - Sisson, Gary

AU - Black, Karen M.

AU - Lin, Liang Tzung

AU - Bilimoria, Darius

AU - Plemper, Richard K.

AU - Privé, Gilbert G.

AU - Richardson, Christopher D.

PY - 2017/12/1

Y1 - 2017/12/1

N2 - The inhibitors carbobenzoxy (Z)-D-Phe-L-Phe-Gly (fusion inhibitor peptide [FIP]) and 4-nitro-2-phenylacetyl amino-benzamide (AS-48) have similar efficacies in blocking membrane fusion and syncytium formation mediated by measles virus (MeV). Other homologues, such as Z-D-Phe, are less effective but may act through the same mechanism. In an attempt to map the site of action of these inhibitors, we generated mutant viruses that were resistant to the inhibitory effects of Z-D-Phe-LPhe- Gly. These 10 mutations were localized to the heptad repeat B (HRB) region of the fusion protein, and no changes were observed in the viral hemagglutinin, which is the receptor attachment protein. Mutations were validated in a luciferase-based membrane fusion assay, using transfected fusion and hemagglutinin expression plasmids or with syncytium-based assays in Vero, Vero-SLAM, and Vero-Nectin 4 cell lines. The changes I452T, D458N, D458G/V459A, N462K, N462H, G464E, and I483R conferred resistance to both FIP and AS-48 without compromising membrane fusion. The inhibitors did not block hemagglutinin protein-mediated binding to the target cell. Edmonston vaccine/laboratory and IC323 wild-type strains were equally affected by the inhibitors. Escape mutations were mapped upon a three-dimensional (3D) structure modeled from the published crystal structure of parainfluenzavirus 5 fusion protein. The most effective mutations were situated in a region located near the base of the globular head and its junction with the alpha-helical stalk of the prefusion protein. We hypothesize that the fusion inhibitors could interfere with the structural changes that occur between the prefusion and postfusion conformations of the fusion protein.

AB - The inhibitors carbobenzoxy (Z)-D-Phe-L-Phe-Gly (fusion inhibitor peptide [FIP]) and 4-nitro-2-phenylacetyl amino-benzamide (AS-48) have similar efficacies in blocking membrane fusion and syncytium formation mediated by measles virus (MeV). Other homologues, such as Z-D-Phe, are less effective but may act through the same mechanism. In an attempt to map the site of action of these inhibitors, we generated mutant viruses that were resistant to the inhibitory effects of Z-D-Phe-LPhe- Gly. These 10 mutations were localized to the heptad repeat B (HRB) region of the fusion protein, and no changes were observed in the viral hemagglutinin, which is the receptor attachment protein. Mutations were validated in a luciferase-based membrane fusion assay, using transfected fusion and hemagglutinin expression plasmids or with syncytium-based assays in Vero, Vero-SLAM, and Vero-Nectin 4 cell lines. The changes I452T, D458N, D458G/V459A, N462K, N462H, G464E, and I483R conferred resistance to both FIP and AS-48 without compromising membrane fusion. The inhibitors did not block hemagglutinin protein-mediated binding to the target cell. Edmonston vaccine/laboratory and IC323 wild-type strains were equally affected by the inhibitors. Escape mutations were mapped upon a three-dimensional (3D) structure modeled from the published crystal structure of parainfluenzavirus 5 fusion protein. The most effective mutations were situated in a region located near the base of the globular head and its junction with the alpha-helical stalk of the prefusion protein. We hypothesize that the fusion inhibitors could interfere with the structural changes that occur between the prefusion and postfusion conformations of the fusion protein.

KW - Antiviral inhibitors

KW - AS-48

KW - Drug resistance

KW - Escape mutations

KW - F protein

KW - FIP

KW - Fusion inhibitors

KW - Measles virus

KW - Membrane fusion

KW - Z-D-Phe-L-Phe-Gly

UR - http://www.scopus.com/inward/record.url?scp=85033785949&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85033785949&partnerID=8YFLogxK

U2 - 10.1128/JVI.01026-17

DO - 10.1128/JVI.01026-17

M3 - Article

C2 - 28904193

AN - SCOPUS:85033785949

VL - 91

JO - Journal of Virology

JF - Journal of Virology

SN - 0022-538X

IS - 23

M1 - e01026-17

ER -