Clark Lab

Department of Biology
University of Texas at Arlington

GRADUATE STUDENTS: Dr. Clark is recruiting graduate students for the Ph.D. program in Quantitative Biology. Please contact Dr. Clark for more information.

Research in my lab focuses on the broad topics of caspase assembly and cell death. Caspases are integral proteases in programmed cell death (apoptosis), and the dysregulation of apoptosis is observed in a number of human diseases, from autoimmune diseases (rheumatoid arthritis, diabetes), to neurodegenerative diseases, to cancer.

About 2.5 million deaths are registered in the US each year (www.cdc.gov). Heart disease and cancer account for about half of the deaths. The impact of chronic diseases (heart disease, stroke, diabetes, Alzheimer) on healthcare costs approaches $750 billion per year. Anticancer drugs are effective at inducing apoptosis by a variety of mechanisms because cancer cells are known to evade pro-apoptotic signals when compared to normal cells. Our goal is to understand how caspase enzyme activity is regulated under normal conditions and how aberrant regulation contributes to disease.

In addition to the cell death programs, caspase enzyme activity is used by cells during development and differentiation in reactions that do not result in cell death. It is not known how cells regulate caspase activity for cell differentiation versus cell death. Levels of active caspases are responsive to cell signaling pathways, and the post-translational modifications of caspases appear to be a key mechanism for fine-tuning activity.

We use biochemical and structural studies to understand the allosteric regulation of caspases and to develop compounds that affect caspase activity through allostery. We have developed an extensive library of allosteric mutants of caspase-3 which allow us to fine-tune caspase activity in cellulo and in animal models.

We study protein structure and function using X-ray crystallography, spectroscopy and other biophysical techniques. We use phage display and other high through-put screening assays to develop drugs for examining caspase allostery, and we examine the regulation of caspase activity in cellular and animal models.

Featured Research:
Identifying Allosteric Modulators through Phage Display

Several sites evolved on caspases that allow binding of allosteric regulators. Some sites are common among caspases while others are unique to an individual caspase. The allosteric sites may provide mechanisms for cells to regulate several caspases simultaneously, using the common sites, or individually, using the unique sites.

We are examining the function and evolution of allosteric sites of caspases using peptide-based phage display technology. This technology rapidly identifies "hot spots" on the protein surface that are important for function. Combinatorial peptide libraries are constructed by inserting DNA encoding random or constrained peptides into gene III of bacteriophage M13. The peptide fusions are displayed on the surface of the bacteriophage and are screened in high-throughput assays to determine binding to a particular caspase. Peptides are then synthesized for in vitro biophysical/biochemical characterization.

Our goal is map the "hot spots" on caspases and to deterimine how cells use the allosteric sites to modify activity in vivo. In diseases with aberrant caspase regulation, such as cancer and neurodegeneration, disrupting the allosteric sites may provide novel therapies to treat the disease.

Publications

A complete list of Dr. Clark's publications can be found on his Google Scholar's Page, and pdf files are posted on his ResearchGate Page. (Both Links Open New Window)

Most Recent:

Modifications to a common phosphorylation network provide individualized control in caspases.
Journal of Biological Chemistry 293, 5447-5461. 2018
Thomas, ME| Grinshpon, R.| Swartz, P. | Clark AC

Tunable allosteric library of caspase-3 identifies coupling between conserved water molecules and conformational selection.
Proceedings of the National Academy of Sciences USA 113, 6080-6088. 2016
Maciag, JJ | MacKenzie SH | Tucker MB | Schipper, JL | Swartz, PD | Clark AC

Phage display and structural studies reveal plasticity in substrate specificity of caspase-3a from zebrafish.
Protein Science 25, 2076-2088. 2016
Tucker, MB | MacKenzie SH | Maciag, JJ | Dirscherl Ackerman, H | Swartz, PD | Yoder, JA | Hamilton, PT | Clark AC

Caspase allostery and conformational selection.
Chemical Reviews 116, 6666-6706. 2016
Clark AC


Caspases - Key Players in Apoptosis,
in Proteases in Apoptosis: Pathways, Protocols and Translational Advances, Bose K. (ed.), Springer, pp 31-51. 2015
Cade C | Clark AC


Modifying caspase-3 activity by altering allosteric networks.
Biochemistry 53: 7582-95. 2014
Cade C | Swartz P | MacKenzie SH | Clark AC |

Keywords

Allostery, Apoptosis, Caspases, Cancer Biology, Protein Engineering, X-ray Crystallography, Spectroscopy (Equilibrium and Kinetic Studies), Isother mal Titration Calorimetry, Analytical Ultracentrifugation, Molecular Modeling, Molecular Dynamics, Site-Directed Mutagenesis, Assay Development, Transfection, Western Analysis, RT-PCR, Cell Sorting, Transgenic Animals, Phage Display