Mair E. A. Churchill, Ph.D.
Associate Professor of Pharmacology
Department of Pharmacology
University of Colorado at Denver and Health Sciences Center
RC1 South Tower, Room 6113
P.O. Box 6508 MS 8303
Aurora, Colorado 80045
Phone: 303.724-3670
Fax: 303.724-3663
Mair.Churchill@uchsc.edu
EDUCATION/EXPERIENCE
AWARDS, MEMBERSHIPS
& FELLOWSHIPS
PROFESSIONAL ACTIVITIES
POSTDOCTORAL POSITIONS
AVAILABLE
Research Interests:
Structural Analysis of Gene-Regulatory Protein
Complexes
My laboratory is interested in the way
that molecular recognition controls protein function in fundamental
molecular processes and in human disease. We use biochemical, biophysical,
and structural tools to study chromosomal proteins, DNA modification
enzymes and enzymes involved in bacterial pathogenesis. Learning more
about structure and mechanism in these systems will advance our understanding
of fundamental molecular processes and aid in the development of novel
pharmaceuticals.
One area of interest is chromatin structure
and function. DNA
in chromatin is packaged and condensed into higher order structures,
but remains accessible to factors involved in processes, such as transcription,
through a complex array of protein-DNA and protein-protein interactions,
as well as protein and DNA modifications. The chromosomal proteins
that bind DNA directly and that are important for the definition of
chromatin structure and regulation of gene expression must be able
to bind to many different DNA sequences. This is in contrast to better
characterized proteins, such as transcription factors, that recognize
a specific sequence of DNA. Histone H1 and the HMG-box proteins are
examples of chromosomal proteins that bind to the linker DNA (between
nucleosomes) and recognize distinct features of DNA structure, such
as shape and flexibility. We are interested in understanding how these
proteins recognize DNA and how these complexes are involved in mediating
cellular processes.
The Drosophila melanogaster HMG-box protein, HMG-D has been the focus
of our study. Through mutagenesis, thermodynamic, and structural analyses,
we have learned how HMG-D binds to DNA non-sequence-specifically (Figure
A), and determined many of the features of the protein that are important
for protein induced DNA bending.
|
| Figure A. We have determined the structure
of the complex of HMG-D bound to linear duplex DNA using X-ray
crystallography. HMG-D severely bends the DNA by binding and
partially intercalating residues in the DNA minor groove. The
structure of this non-sequence- specific protein-DNA complex
is similar to homologous sequence-specific complexes, except
for the lack of sequence-specific hydrogen bonds. Instead, hydrophobic
interactions and water mediated non-specific hydrogen bonds
stabilize the complex. |
Our work on the non-sequence-specific HMG-box proteins has contributed
to understanding how abundant chromosomal proteins interact with DNA
and how they may influence the behavior of other protein-DNA complexes.
Future studies will focus on testing these hypotheses through structural
and biophysical analyses of multi-protein-DNA complexes important
in gene regulation.
A second area of interest is in the mechanism
of DNA modification. The RsrI methyltransferase (M.RsrI) is
a component of the Rhodobacter sphaeroides restriction-modifcation
system that protects bacteria from invasion by bacteriophages and
foreign DNA. MoRsrI methylates the N6 position of adenine within the
recognition site GAATTC. In collaboration with Dr. Richard Gumport's
group at the U. of Illinois at Urbana-Champaign, we have determined
the structure of M.RsrI, which suggests a novel mechanism of DNA binding
for the methyltransferases (see Figure B). Future studies include
analysis of M.RsrI interactions with DNA, and structure determination
of other methyltransferases and demethylases important in modulation
of DNA structure.
|
Figure B. Structure of
M.RsrI: The structure explains how DNA recognition and methylation
may occur, when the required functional domains reside on opposite
sides of the enzyme monomer. A unique dimer of the enzyme is
observed in the crystal. This configuration brings the DNA binding
domain of one subunit (aqua) near the enzyme active site, which
contains a cofactor analog, 5'methylthioadenosine, bound to
the red monomer on the right. |
A third research interest is global gene
regulation in bacteria. As we enter the post-antibiotic era,
it is more important now than ever before to understand the molecular
and structural basis of bacterial pathogenicity. Quorum sensing, the ability of the bacteria to sense their
local concentration, regulates bacterial pathogenicity by altering
gene expression on a global scale.
Quorum sensing in gram negative bacteria depends on a simple
lipid mediator called acyl-homoserinelactone (AHL) that is synthesized
by the AHL-synthase, and is detected by a response regulator transcription
factor. We are studying
the quorum sensing systems in several pathogenic gram negative bacteria
to understand the mechanistic basis for AHL synthesis and specificity. Our structural studies provide the foundation
for the development of pharmacological agents for treatment of persistent
as well as multi-drug resistant forms of bacterial infection.
Selected Publications
-
M.-H. Kuo, J. Zhou, P. Jambeck, M.E.A.
Churchill and C. D. Allis, "Histone Acetyltransferase
Activity of Yeast Gcn5p is Required for the Activation of Target
Genes in vivo" (1998) Genes Dev., 12, 627-639. [Abstract]
-
Churchill, M.E.A., Changela,
A., Dow, L.K. and Krieg, A.J., "Interactions of HMG-box
proteins with DNA and chromatin" (1999) Methods in Enzymology
304, 99-131. [Abstract]
-
F.V. Murphy IV, R. M. Sweet, and M.E.A.
Churchill "The Structure of a Chromosomal High-Mobility-Group
Protein-DNA Complex Reveals Sequence-neutral Mechanisms Important
for Non-sequence-specific DNA Recognition." (1999) EMBO
J. 18, 6610-6618. [Abstract]
-
F.V. Murphy IV and M.E.A. Churchill
"Non-sequence-specific DNA recognition: a structural perspective."
(2000) Structure 8, R83-89. [Abstract]
-
Dow LK, Jones DN, Wolfe SA, Verdine GL, Churchill
ME. Structural studies of the high mobility group globular
domain and basic tail of HMG-D bound to disulfide cross-linked
DNA. Biochemistry. 2000 Aug 15;39(32):9725-36. [Abstract]
-
Scavetta RD, Thomas CB, Walsh MA, Szegedi S, Joachimiak A,
Gumport RI, Churchill ME.
Structure of RsrI methyltransferase, a member of the N6-adenine
beta class of DNA methyltransferases. Nucleic Acids Res. 2000
Oct 15;28(20):3950-61. [Abstract]
- Watson WT, Minogue TD, Val DL, von Bodman SB, Churchill ME.
Structural basis and specificity of acyl-homoserine lactone signal
production in bacterial quorum sensing. Mol Cell 2002 Mar;9(3):685-94
[Abstract]
- Melvin VS, Roemer SC, Churchill ME, Edwards DP.
The C-terminal extension (CTE) of the nuclear hormone receptor
DNA binding domain determines interactions and functional response
to the HMGB-1/-2 co- regulatory proteins.
J Biol Chem 2002 Jul 12;277(28):25115-24 [Abstract]
Bibliography
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