The long-term goal of the research described in this application is to find ways to improve vaccines against epidemic influenza in the human population. The objective of the proposed experiments is to gain better understanding of how antibodies neutralize virus infectivity and the mechanisms by which influenza A viruses escape from neutralizing antibodies. We will focus effort on the viral neuraminidase (NA) since this is the only viral antigen for which a crystal structure of an antigen-antibody complex is available. Also antibody inhibition of NA enzyme activity can be quantitatively measured. However, we have not forgotten that the hemagglutinin (HA) is more plentiful on the virus surface, has better-defined and multiple functions, and that anti-HA antibodies directly neutralize infectivity. Therefore we propose experiments to use the procedures we will have worked out for the NA to investigate mechanisms of neutralization by anti-HA antibodies.
The Specific Aims are to answer the following questions: 1. How does the influenza neuraminidase escape from antibody inhibition? Site-directed mutagenesis procedures will be used to test the hypothesis that only some of the 22 amino acids on subtype N9 NA which make contact with antibody NC41 are essential to the interaction, and therefore that only certain changes can lead to antigenic drift. 2. Are the principles of virus escape deduced from N9 NA relevant to human influenza? The antigenic structure and variation of N2 NA from a recent human isolate, A/Shanghai/11/87 will be studied using monoclonal antibodies, following procedures developed for the avian virus N9 NA which have been validated by x-ray crystallographic results. 3. What is the mechanism of inhibition by anti-NA antibodies such as NC41? What function of influenza virus NA are they inhibiting? The x-ray crystal structure of NA-antibody complex suggests that inhibition is not necessarily stearic. The effect of antibody on the NA active site will be determined, and the role of NA as a receptor-destroying enzyme will be investigated. 4. Mechanism of neutralization by anti-HA antibodies. Antibodies against the HA are classical neutralizing antibodies, unlike those to the NA which act after infection has begun. We will apply the procedures developed for studying antibody inhibition of NA to localize neutralizing epitopes on a recent human HA.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37AI019084-11
Application #
3481005
Study Section
Experimental Virology Study Section (EVR)
Project Start
1983-04-01
Project End
1996-03-31
Budget Start
1993-04-01
Budget End
1994-03-31
Support Year
11
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of Alabama Birmingham
Department
Type
Schools of Dentistry
DUNS #
004514360
City
Birmingham
State
AL
Country
United States
Zip Code
35294
Lee, Janis T; Air, Gillian M (2002) Contacts between influenza virus N9 neuraminidase and monoclonal antibody NC10. Virology 300:255-68
Gulati, Upma; Hwang, Chi-Ching; Venkatramani, Lalitha et al. (2002) Antibody epitopes on the neuraminidase of a recent H3N2 influenza virus (A/Memphis/31/98). J Virol 76:12274-80
Ghate, A A; Air, G M (1999) Influenza type B neuraminidase can replace the function of type A neuraminidase. Virology 264:265-77
Pruett, P S; Air, G M (1998) Critical interactions in binding antibody NC41 to influenza N9 neuraminidase: amino acid contacts on the antibody heavy chain. Biochemistry 37:10660-70
Liu, C; Eichelberger, M C; Compans, R W et al. (1995) Influenza type A virus neuraminidase does not play a role in viral entry, replication, assembly, or budding. J Virol 69:1099-106
Nuss, J M; Air, G M (1994) Defining the requirements for an antibody epitope on influenza virus neuraminidase: how tolerant are protein epitopes? J Mol Biol 235:747-59
Zhang, H; Air, G M (1994) Expression of functional influenza virus A polymerase proteins and template from cloned cDNAS in recombinant vaccinia virus infected cells. Biochem Biophys Res Commun 200:95-101
Nuss, J M; Whitaker, P B; Air, G M (1993) Identification of critical contact residues in the NC41 epitope of a subtype N9 influenza virus neuraminidase. Proteins 15:121-32
Nuss, J M; Air, G M (1991) Transfer of the hemagglutinin activity of influenza virus neuraminidase subtype N9 into an N2 neuraminidase background. Virology 183:496-504
Air, G M; Laver, W G; Webster, R G (1990) Mechanism of antigenic variation in an individual epitope on influenza virus N9 neuraminidase. J Virol 64:5797-803

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