Chemistry of Cellular Regulation 
Paul B. Sigler, M.D., Ph.D. — Investigator 
Dr. Sigler is also Professor of Molecular Biophysics and Biochemistry at Yale University. He studied 
chemistry at Princeton University and received his M.D. degree from Columbia University. He then spent 
two years as a house officer in the Department of Medicine at Columbia-Presbyterian Medical Center, 
New York. He began his work on crystallography with David Davies at NIH. He studied as a Helen Hay 
Whitney Fellow at the MRC Laboratory of Molecular Biology in Cambridge, England, where he received 
his Ph.D. degree in biochemistry. Before accepting his present position. Dr. Sigler was Professor of 
Biochemistry and Molecular Biology at the University of Chicago. Dr. Sigler was recently elected to the 
National Academy of Sciences. 
MY colleagues and I continue to pursue our 
interest in the molecular mechanism of reg- 
ulation at the level of transcription and trans- 
membrane signaling. Our approach is to use 
high-resolution x-ray crystallography to deter- 
mine the structure of the relevant macromole- 
cules and their assemblies. We infer chemical 
mechanisms from these structures and test them 
with biochemical and physicochemical experi- 
ments and with directed mutagenesis. 
Transcriptional Regulation 
In the area of transcriptional control, our pri- 
mary focus remains the mechanism by which 
DNA-binding regulatory proteins are targeted to 
their responsive genes. Our earliest and best- 
studied experimental system has been the alloste- 
ric regulation of the trp repressor and the forma- 
tion of the trp repressor-operator complex. 
Having defined the stereochemistry of the 
protein-DNA interface, we are now extending the 
study (partly in collaboration with Julian Sturte- 
vant, Yale University) to include an analysis of 
the thermodynamic parameters underlying the 
trp repressor's ability to bind selectively to the 
trp operator. 
Crystallographic structural analyses have been 
extended to the mechanism by which steroid re- 
ceptors bind selectively to their DNA targets. The 
first complex in this series (in collaboration with 
Len Freedman and Keith Yamamoto, University 
of California, San Francisco) shows the DNA- 
binding domain in complex with its idealized tar- 
get, the symmetrical glucocorticoid response ele- 
ment. It shows how the two zinc fingers of the 
domain interact with each other to form a unified 
globular domain and how the domain makes 
chemical contacts with the bases that identify the 
"half sites" of the symmetrical response element. 
It also shows a surprisingly novel mechanism by 
which the receptor recognizes the invariant 
three-base pair spacing between the half sites. 
Interactions with the DNA phosphates stabilize 
a change in the protein's structure that causes it 
to form a firm dimer. In so doing, the DNA-bind- 
ing surfaces of each subunit are placed in perfect 
registry with the complementary surfaces of the 
DNA target's half sites. Thus the DNA itself helps 
form a dimer that discriminates between DNA se- 
quences that have the correct spacings between 
their half sites and those that do not. These same 
DNA-induced conformational changes are also 
likely to potentiate functional contacts between 
the receptor and other components of the tran- 
scription initiation assembly. 
Work is under way to examine the arrangement 
by which the glucocorticoid receptor contacts 
other transcription factors on "composite" regu- 
latory sites. We are also examining the way other 
members of this steroid receptor superfamily in- 
teract with their respective response elements. 
Only through a comparative study can we build a 
reliable picture of the targeting mechanism. 
Most recently we have solved (in collaboration 
with Laimonis Laimins, HHMI, University of Chi- 
cago) the structure of the complex between the 
E2 transcription factor of bovine papilloma virus 
and the DNA sequence (enhancer) to which it 
binds. E2 is involved in replication of the viral 
genome and controls expression of the genes re- 
sponsible for transformation of cells infected 
with this oncogenic virus. The structure has been 
refined at an unprecedented level of detail (1.7 
A) and exhibits a barrel-like dimeric architecture 
that has never been seen before in any protein 
structure, let alone in a DNA-binding domain. 
The DNA is severely bent over this barrel as it 
forms specific contacts with the protein. These 
results are now being correlated with genetic in- 
formation produced by others. 
Several other specific DNA complexes of tran- 
scriptional regulatory proteins are in earlier 
stages of crystallographic analysis. A particularly 
interesting one is the arg repressor, which regu- 
lates the expression of arginine biosynthetic 
genes. It is unique because it is a hexamer of 
identical subunits. Its target is also unusual, in 
that it is two symmetrical 1 8-base pair sequences 
whose symmetry axes are separated by 2 1 base 
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