# IMMUNOCHEMICAL STUDIES OF HUMAN RESPONSE TO ALLERGENS

> **NIH NIH R01** · JOHNS HOPKINS UNIVERSITY · 1992 · $253,743

## Abstract

The long-term goal of this research is to understand the immunochemical
basis of human immune responsiveness to inhaled pollen allergens.  We will
study two homologous groups of ragweed allergenic proteins, the Amb Vs and
the Amb VIs.  Responsiveness to the Amb V homologues is known to be
associated with HLA-DR2/Dw2 (DR2.2), and DR2.12 in Orientals;
responsiveness to Amb VI is associated with DR5 (DRw11) plus DR2.21 (and
probably also DR2.22).  Interestingly, responsiveness to Amb a VI is
negatively associated with DR2.2.  Within a homologous group, we postulate
that the portions of these molecules (agretopes) that bind to the Class II
MHC (Ia) molecules are similar (accounting for the same Ia restriction),
but that the T-cell and B-cell epitopes are usually different for the
individual homologues.  Analysis of these epitopes will be facilitated by
use of NMR analysis and computer modeling.

We will sequence Amb V cDNAs of several ragweed species.  Protein
structures of Amb a V and Amb t V will be determined by NMR spectroscopy,
and the structures of further Amb Vs by NMR or computer modeling.
Functional, recombinant Amb a V, Amb t V and Amb p V proteins will be
produced by cloning the respective cDNAs into the pGEX-2T expression vector
in E. coli.  Mutations that are potentially informative in defining Ia/T-
cell and B-cell epitopes will be identified from the NMR structures and by
computer modeling.  We will then produce the Amb V variants by site-
directed mutagenesis and expression of the cDNAs.  The Ia-binding and T-
cell-binding portions of the Ia/Tcell epitopes, as well as the B-cell
epitopes of the Amb V molecules will be defined by examining these mutants,
as well as using synthetic peptides (native and variant sequences) of Amb
Vs.  Mutant Amb Vs that have altered immunological properties will be
analyzed by NMR to examine the effects of altering selected amino acids.
Since the Amb Vs have a complex array of disulfide bonds that are
implicated in T-cell epitopes, these studies will provide a valuable model
for the molecular analysis of Ia/T-cell epitopes containing disulfide(s).
We will transfect HLA-DRA and DRB1*1501 (encoding DRbetaI2.2), and mutants
of the betaI gene, into a human B-cell line that does not express Ia or
into mouse L cells.  Using these constructs, we will determine which Ia
residues are critical for binding to Amb V peptides.

Similar studies, including cDNA cloning and sequencing, will be performed
in the case of the Amb VIs; in this case, it will also be interesting to
examine the structural basis for why the immune responsiveness is
positively associated with DR5/DR2.21, but negatively associated with
DR2.2.

A unique asset will be the availability of two well-characterized
populations:  ca 400 clinic patients and ca 500 subjects from an
epidemiologic-genetic study of human immune response to allergens.
Subjects from different racial and ethnic origins, included in an
international HLA and Allergy project, will al...

## Key facts

- **NIH application ID:** 3129101
- **Project number:** 2R01AI019727-09A1
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **SAM.gov UEI:** FTMTDMBR29C7
- **Principal Investigator:** DAVID GEORGE MARSH
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 1992
- **Award amount:** $253,743
- **Award type:** 2
- **Project period:** 1983-08-01 → 1997-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/3129101

## Citation

> US National Institutes of Health, RePORTER application 3129101, IMMUNOCHEMICAL STUDIES OF HUMAN RESPONSE TO ALLERGENS (2R01AI019727-09A1). Retrieved via MORVS 2026-10-25 from https://api.morvs.ai/grant/nih/3129101. Licensed CC0.

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