Localization of monoclonal antibody epitopes and functional domains in the hemagglutinin protein of measles virus

Hummel, Kimberly B.; Bellini, William J.

doi:10.1128/jvi.69.3.1913-1916.1995

Cited by 38 publications

(6 citation statements)

References 33 publications

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“…Consistent with results of previous studies (12), the genetic sequence of the entire H gene of five randomly chosen MAbresistant mutants confirmed that a single point mutation is sufficient to confer the resistance phenotype. The sequence analysis revealed, however, that these single point mutations did not all occur at the same site within the H gene.…”

Section: Discussionsupporting

confidence: 89%

Spontaneous Mutation Rate of Measles Virus: Direct Estimation Based on Mutations Conferring Monoclonal Antibody Resistance

Schrag

Rota

Bellini

1999

J Virol

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High mutation rates typical of RNA viruses often generate a unique viral population structure consisting of a large number of genetic microvariants. In the case of viral pathogens, this can result in rapid evolution of antiviral resistance or vaccine-escape mutants. We determined a direct estimate of the mutation rate of measles virus, the next likely target for global elimination following poliovirus. In a laboratory tissue culture system, we used the fluctuation test method of estimating mutation rate, which involves screening a large number of independent populations initiated by a small number of viruses each for the presence or absence of a particular single point mutation. The mutation we focused on, which can be screened for phenotypically, confers resistance to a monoclonal antibody (MAb 80-III-B2). The entire H gene of a subset of mutants was sequenced to verify that the resistance phenotype was associated with single point mutations. The epitope conferring MAb resistance was further characterized by Western blot analysis. Based on this approach, measles virus was estimated to have a mutation rate of 9 × 10−5 per base per replication and a genomic mutation rate of 1.43 per replication. The mutation rates we estimated for measles virus are comparable to recent in vitro estimates for both poliovirus and vesicular stomatitis virus. In the field, however, measles virus shows marked genetic stability. We briefly discuss the evolutionary implications of these results.

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Section: Discussionsupporting

confidence: 89%

Spontaneous Mutation Rate of Measles Virus: Direct Estimation Based on Mutations Conferring Monoclonal Antibody Resistance

Schrag

Rota

Bellini

1999

J Virol

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“…Residues 451 and 481 fall within the region between amino acids 451 and 505 that Hummel and Bellini have suggested forms part of the hemadsorption and hemagglutination domain of the MV H (8). It would appear from our results (Table 1) that residue 481 has the major influence.…”

Section: Discussionsupporting

confidence: 47%

Identification of two amino acids in the hemagglutinin glycoprotein of measles virus (MV) that govern hemadsorption, HeLa cell fusion, and CD46 downregulation: phenotypic markers that differentiate vaccine and wild-type MV strains

Lecouturier

Fayolle

Caballero

et al. 1996

J Virol

159

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We have used site-directed mutagenesis of the hemagglutinin (H) glycoprotein of measles virus (MV) to investigate the molecular basis for the phenotypic differences observed between MV vaccine strains and recently isolated wild-type MV strains. The former downregulate CD46, the putative cellular receptor of MV, are positive for hemadsorption, and are fusogenic in HeLa cells, whereas the latter are negative for these phenotypic markers. CD46 downregulation in particular, could have profound consequences for the immunopathology of MV infection, as this molecule protects the cell from complement lysis. Mutagenesis of two amino acids, valine and tyrosine at positions 451 and 481, respectively, in the H protein from the vaccine-like Hallé MV strain to their counterparts, glutamate and asparagine, in the H protein from the wild-type Ma93F MV strain (creating the V451E/Y481N double mutation) abrogated CD46 downregulation, HeLa cell fusion, and hemadsorption. The converse double mutagenesis of the Ma93F H protein (E451V/N481Y) transferred the CD46-downregulating, fusogenic, and hemadsorption functions to this protein. The data provide the first mapping study of the functional domains of MV H. The consequences of these results for MV vaccine design and the role of CD46 in MV infection are discussed.

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“…Previous competitive enzyme-linked immunosorbent assay (ELISA) showed that the mAb 2F4 did not compete for MV binding with the mAbs, including E81, E185, E39, and E103, which are recognizing epitopes I, iv, v, and vi, respectively [13]. On the other hand, previously reported mAbs, such as B2, BH26, I-41, 80-II-B2, and 16-DE6, similarly or partially recognize the same epitope vii [11,12,[26][27][28], which appears to compete with 2F4. Therefore, the use of 2F4 in combination with the mAbs which bind to distinct epitopes would be more effective for measles neutralization.…”

Section: Discussionmentioning

confidence: 96%

Biophysical characterization and single‐chain Fv construction of a neutralizing antibody to measles virus

et al. 2019

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The measles virus (MV) is a major cause of childhood morbidity and mortality worldwide. We previously established a mouse monoclonal antibody, 2F4, which shows high neutralizing titers against eight different genotypes of MV. However, the molecular basis for the neutralizing activity of the 2F4 antibody remains incompletely understood. Here, we have evaluated the binding characteristics of a Fab fragment of the 2F4 antibody. Using the MV infectious assay, we demonstrated that 2F4 Fab inhibits viral entry via either of two cellular receptors, SLAM and Nectin4. Surface plasmon resonance (SPR) analysis of recombinant proteins indicated that 2F4 Fab interacts with MV hemagglutinin (MV‐H) with a KD value at the nm level. Furthermore, we designed a single‐chain Fv fragment of 2F4 antibody as another potential biopharmaceutical to target measles. The stable 2F4 scFv was successfully prepared by the refolding method and shown to interact with MV‐H at the μm level. Like 2F4 Fab, scFv inhibited receptor binding and viral entry. This indicates that 2F4 mAb uses the receptor‐binding site and/or a neighboring region as an epitope with high affinity. These results provide insight into the neutralizing activity and potential therapeutic use of antibody fragments for MV infection.

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Localization of monoclonal antibody epitopes and functional domains in the hemagglutinin protein of measles virus

Cited by 38 publications

References 33 publications

Spontaneous Mutation Rate of Measles Virus: Direct Estimation Based on Mutations Conferring Monoclonal Antibody Resistance

Spontaneous Mutation Rate of Measles Virus: Direct Estimation Based on Mutations Conferring Monoclonal Antibody Resistance

Identification of two amino acids in the hemagglutinin glycoprotein of measles virus (MV) that govern hemadsorption, HeLa cell fusion, and CD46 downregulation: phenotypic markers that differentiate vaccine and wild-type MV strains

Biophysical characterization and single‐chain Fv construction of a neutralizing antibody to measles virus

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