Protein Structures, Molecular Recognitions, and Functions 
Structure and Recognition of Epitopes 
Knowledge of the active conformation of a 
dominant neutralizing epitope (antigenic deter- 
minant group) is essential to understanding the 
molecular basis of the immunogenicity of the 
segments and to obtaining possible leads for vac- 
cine design. Recombinant technology is being 
used to insert, without loss of immunogenicity, 
foreign epitopes, especially of proteins with un- 
known structures, onto the surface of the malto- 
dextrin-binding protein of Escherichia coli, in 
sites known to be permissive to changes. The 
structure of the protein containing such an epi- 
tope provides information about the epitope's 
configuration and how its immunogenicity is pre- 
served. Currently under investigation are epi- 
topes of type 1 poliovirus and HIV (human immu- 
nodeficiency virus) gpl20, the coat protein of 
the AIDS virus. 
Antibody- Antigen Interaction 
We have previously reported the crystalliza- 
tion of the Fab fragment of a monoclonal anti- 
body raised against a dominant neutralizing deter- 
minant of gpl20. Diffraction data from these 
crystals, out to 2.8 A resolution, have been col- 
lected by means of a synchrotron facility. At- 
tempts to determine the structure, using molecu- 
lar replacement techniques, are under way. X-ray 
structure determination should reveal details of 
molecular recognition of an antigen and impor- 
tant facets of gpl20 that are amenable to anti- 
body intervention. 
We have determined the structure of the Fab 
fragment of an antibody against cell surface poly- 
saccharide antigen of Shigella flexneri in com- 
plex with tri- and pentasaccharide-containing de- 
terminants of the O-antigen serotype of the 
bacteria. Determination of the structures of these 
crystals is in progress. 
Molecular Recognition of Carbohydrates, 
Charged Substrates, and Calcium 
The family of binding proteins that serve as ini- 
tial receptors for bacterial active transport and 
chemotaxis is an excellent system in which to 
study the molecular recognition of a variety of 
ligands and structure-function relationships. To 
date, we have determined and refined at better 
than 1 .9 A resolution the structures of seven dif- 
ferent periplasmic receptor proteins: those 
binding L-arabinose, D-galactose/o-glucose, mal- 
todextrin, leucine/isoleucine/valine, leucine 
alone, sulfate, and phosphate. Using site-directed 
mutagenesis, coupled with analysis of substrate 
binding to mutant proteins by crystallographic 
and other techniques, we are beginning to dissect 
various aspects of the molecular recognition of 
carbohydrates, charged ligands such as sulfate 
and phosphate, and calcium ion. 
A method different from site-directed muta- 
genesis for probing the contribution of specific 
hydrogen bonds in protein-carbohydrate interac- 
tions involves specific modifications of the li- 
gand. The binding of deoxy and fluorodeoxy su- 
gar analogues to the arabinose-binding protein 
has been analyzed by equilibrium and very high 
resolution crystallographic techniques. The com- 
plexes have structures virtually identical to the 
wild type and reflect more realistically the hydro- 
gen bond interaction being probed. 
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