Mechanisms of the Biological Activities of 
Membrane Glycoproteins 
Don C. Wiley, Ph.D. — Investigator 
Dr. Wiley is also Professor of Biochemistry and Biophysics at Harvard University and Research Associate in 
Medicine at the Laboratory of Molecular Medicine at the Children's Hospital, Boston. He received his Ph.D. 
degree in biophysics from Harvard University. He then joined the faculty at Harvard and served as 
Assistant and Associate Professor of Biochemistry and Molecular Biology before attaining his present 
position. Dr. Wiley is Chairman of the Department of Biochemistry and Molecular Biology. He is a member 
of the National Academy of Sciences and a Fellow of the American Academy of Arts and Sciences. Among 
his honors is the Louisa Gross Horwitz Prize from Columbia University. 
Tcell recognition occurs when cell surface his- 
tocompatibility glycoproteins present anti- 
gens, processed to small peptides, to an antibody- 
like molecule on the T cell receptor. An individ- 
ual organism has only a small number of different 
histocompatibility molecules (probably less than 
a dozen), so that each histocompatibility gly- 
coprotein must be able to "present" many, 
possibly thousands, of different antigenic pep- 
tides to thousands or more distinct T cell re- 
ceptors throughout the immunological life of 
the individual. 
In the past year we have been able to visualize 
the conformation of peptides bound to a histo- 
compatibility glycoprotein, HIA-B27. The ex- 
tended conformation of the peptide appears to be 
specified by the HLA-binding site, so that the two 
ends of the peptide are bound to specialized re- 
gions at the two ends of the binding groove. A few 
of the side chains of a 9-mer peptide interact with 
pockets in the surface of the HLA molecule, 
whose size and chemical composition vary from 
allele to allele in the population. We also eluted, 
sequenced, and identified 1 1 self-peptides from 
HLA-B27. All 11 had arginine, a positively 
charged amino acid, at peptide position 2, which 
correlates with the x-ray crystallographic finding 
that position 2 fits into a deep pocket with a nega- 
tively charged glutamic acid at the bottom. (That 
residue is polymorphic and changes the specific- 
ity of that pocket in other alleles.) 
We are also now able to reconstitute class I mol- 
ecules from polypeptide chains produced in 
Escherichia coli with single peptides and have 
crystallized HLA-A2 with a series of peptide anti- 
gens from influenza virus and HIV-l (human im- 
munodeficiency virus type 1 ) . One complex dif- 
fracts beyond 1 .5-A resolution when the crystal is 
frozen at - 160°C. (All of the crystals in our labo- 
ratory are now frozen to this temperature to pre- 
serve crystallographic order and eliminate radia- 
tion damage.) 
In our studies of class II histocompatibility an- 
tigens (a collaboration with Joan Gorga and Jack 
Strominger), we now have three crystals under 
study: human DRl, human DRl plus a superanti- 
gen, and DRl expressed in insect cells and com- 
plexed with a single influenza virus peptide. The 
complex of a single peptide with a class II mole- 
cule was generated by expressing a soluble class 
II molecule in cells from insects, which lack an 
immune system. Empty DRl molecules were pro- 
duced that rapidly and stoichiometrically bound 
peptide. The empty molecules were stabilized 
against aggregation and sodium dodecyl sulfate- 
induced denaturation by addition of peptide, ar- 
guing that peptide binding is accompanied by a 
conformational change. 
Our laboratory is also studying how influenza 
virus infects cells. About 10 years ago we deter- 
mined the three-dimensional structure of the in- 
fluenza virus hemagglutinin (HA), the viral gly- 
coprotein responsible for binding the virus to 
cells and for fusing the viral membrane to a cellu- 
lar membrane to effect infectious entry. Recently 
we determined the structure of a series of com- 
plexes between the HA and derivatives of sialic 
acid, the cellular receptor for influenza virus. We 
have synthesized a number of these new ligands 
and determined the crystal structure of the com- 
plexes to confirm an atomic model for virus-cell 
binding that we proposed. In the process a sec- 
ond binding site has been located on the HA at an 
interface between domains of the molecule, 
which, although probably not physiological, may 
offer opportunities for the design of a ligand to 
stabilize the interface against the conformational 
change required for the HA's membrane fusion 
activity. 
A number of other crystallographic and bio- 
chemical studies are under way on influenza C 
virus, on a low-pH fusion-active conformation of 
the influenza HA, on trypanosome surface anti- 
gens, and on the glycoprotein of HIV- 1 in com- 
plex with its cellular receptor, CD4. 
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