Structural Studies of Macromolecular Assemblies 
nents of a transcriptional initiation complex pres- 
ent even more challenging puzzles for the future. 
Viruses 
The small, double-stranded DNA viruses SV40 
(simian virus 40) and polyoma have given us a 
first glimpse of virus particles that package a mini- 
chromosome in one cell and deliver it to the nu- 
cleus in another. The shells of these viruses are 
composed of 72 pentamers of the major struc- 
tural protein VPl and 30-60 copies each of two 
internal proteins, VP2 and VP3. These compo- 
nents package the viral DNA. The VPl polypep- 
tide chain folds in such a w^ay that two large 
j8-sheets with radially directed strands form a 
framework, with very tight interactions between 
adjacent subunits in a pentamer. The carboxyl 
terminus of VPl forms an extended arm that in- 
teracts with subunits of another pentamer, gener- 
ating three kinds of interpentamer contact in the 
virus particle. This tying together of standard 
building blocks allows for the required variabil- 
ity in packing geometry without sacrificing speci- 
ficity. Flexibly extended arms, which form or- 
dered structures only when the units assemble 
into a particle, appear to be an important feature 
of complex assemblies. 
A recently determined structure for the murine 
polyoma virus shows that an important surface 
loop is larger in the polyoma subunit than in 
SV40. Mutational evidence suggests that this loop 
creates a shallow pocket for binding sialic acid, 
required for cell entry by polyoma but not by 
SV40. A number of viruses of various structural 
types use cell-surface sialic acid for attachment, 
and it is of broad interest to understand how spe- 
cific viruses accomplish the interaction. Compar- 
ison of SV40 and polyoma shows that this func- 
tion can readily be added or lost by small changes 
at the surface of a viral coat protein. 
We have recently begun to study the double- 
stranded RNA viruses. Crystals of rotavirus single- 
shelled particles and reovirus cores diflfract to 
at least 7-A resolution. These particles are elab- 
orately organized molecular machines, con- 
taining the complete transcription and RNA- 
modification activities. 
Receptors 
The receptor for human immunodeficiency 
virus (HIV) is the lymphocyte surface antigen 
CD4. Its extracellular portion is composed of 
four immunoglobulin-like domains. We have de- 
termined the atomic structure of a two-domain, 
amino-terminal fragment, which binds HIV as 
tightly as does the intact receptor. The first two 
domains are joined by a continuous (8-strand con- 
nector, and they have an extensive hydrophobic 
interface. Thus they form a rigid, rod-like seg- 
ment. The HIV-binding site appears to be a ridge 
along one edge of the first domain. Binding and 
mutational studies, carried out collaboratively, 
show that a projecting phenylalanyl side chain is 
critical for the interaction with HIV gpl20 and 
that various positively charged residues sur- 
rounding it are also important. The same region 
appears to be involved in contacts with class II 
MHC (major histocompatibility complex) mole- 
cules. We have also crystallized a four-domain 
fragment of CD4, corresponding to the entire ex- 
tracellular region, and the structure determina- 
tion is in progress. 
Many important receptors are taken up into 
the cell by a process of endocytosis mediated 
by clathrin-coated vesicles. The transferrin re- 
ceptor is one such molecule. It undergoes a well- 
characterized cycle of uptake and return to the 
cell surface. We have crystallized the extracellu- 
lar domain, which makes up about three-fourths 
of the molecule. This domain exhibits reversible 
conformational changes at low pH that we be- 
lieve to be significant for intracellular sorting 
steps. The determination of the structure has re- 
cently been facilitated by the observation that 
much better diffraction data can be collected by 
studying crystals at liquid-nitrogen temperatures. 
Such freezing techniques are now being broadly 
applied in our laboratory, enhancing our ability 
to study radiation-sensitive crystals of very com- 
plex structures. 
178 
