Scientific Overview: Structural Sciences

Research Interests

Our laboratory is employing multidisciplinary approaches to study the structures and functions of protein complexes that play key roles in major signaling pathways, and to use the resulting structural information to develop therapeutic agents for the treatment of human disease, including cancer and diabetes. Currently we are focusing on three families of proteins: nuclear hormone receptors, the Met tyrosine kinase receptor, and G protein–coupled receptors, because these proteins, beyond their fundamental roles in biology, are important drug targets for many human diseases.

Nuclear hormone receptors

The nuclear hormone receptors form a large family comprising ligand-regulated and DNA-binding transcriptional factors. The family includes receptors for classic steroid hormones such as estrogen, progesterone, androgens, and glucocorticoids, as well as receptors for peroxisome proliferator activators, vitamin D, vitamin A, and thyroid hormones. One distinguishing fact about these classic receptors is that they are among the most successful targets in the history of drug discovery: every receptor has one or more cognate synthetic ligands currently being used as medicines. The nuclear receptors also include a class of “orphan” receptors for which no ligand has been identified. In the last two years, we have developed the following projects centering on the structural biology of nuclear receptors.

Peroxisome proliferator–activated receptors

The peroxisome proliferator–activated receptors (PPARα, δ, and γ) are key regulators of glucose and fatty acid homeostasis and as such are important therapeutic targets for treating cardiovascular disease, diabetes, and cancer. To understand the molecular basis of ligand-mediated signaling by PPARs, we have determined crystal structures of each PPAR’s ligand-binding domain (LBD) bound to diverse ligands including fatty acids, the lipid-lowering fibrate drugs, and a new generation of anti-diabetic drugs, the glitazones. We have also determined the crystal structures of these receptors bound to coactivators or co-repressors. We are developing this project into the structures of large PPAR fragment/DNA complexes.

Human glucocorticoid and mineralocorticoid receptors

The human glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) are classic steroid hormone receptors that are key to a wide spectrum of human physiology including immune/inflammatory responses, metabolic homeostasis, and control of blood pressure. Both are well-established drug targets. GR ligands such as dexamethasone (Dex) and fluticasone propionate (FP) are used to treat asthma, leukemia, and autoimmune diseases; MR ligands such as spironolactone and eplerenone are used to treat hypertension and heart failure. However, the clinical use of these ligands is limited by undesirable side effects partly associated with their receptor cross-reactivity or low potency. Thus, the discovery of highly potent and more-selective ligands for GR and MR is an important goal of pharmaceutical research.

We have determined a crystal structure of the GR LBD bound to dexamethasone and the MR LBD bound to corticosterone, both of which are in complex with a coactivator peptide motif. These structures provide a detailed basis for the specificity of hormone recognition and coactivator assembly by GR and MR. Currently we are studying receptor-ligand interactions by crystallizing GR and MR with various steroid or nonsteroid ligands. In collaboration with Brad Thompson and Raj Kumar at the University of Texas Medical Branch at Galveston, we are also extending our studies to the structure of a large GR fragment bound to DNA.

The human androgen receptor

The androgen receptor (AR) is the central molecule in the development and progression of prostate cancer, and as such it serves as the molecular target of anti-androgen therapy. However, most prostate cancer patients develop resistance to such therapy, mainly due to mutations in this hormone receptor that alter its three-dimensional structure and allow AR to escape repression. The growth of prostate cancer cells that harbor a mutated AR is then no longer dependent on androgen, making anti-hormone therapy ineffective. This form of hormone-independent prostate cancer is highly aggressive and is responsible for most deaths from prostate cancer. The development of effective therapies requires a detailed understanding of the structure and functions of the central molecule, i.e., the androgen receptor and its interactions with hormones and co-regulators. In this project, we are aiming to determine the structures of the mutated AR proteins that alter the response to anti-hormone therapy. In collaboration with Donald MacDonnell at Duke University, we are working on the crystal structure of the full-length AR/DNA complex.

Structural genomics of nuclear receptor ligand-binding domains

The LBDs of nuclear receptors contain key structural elements that mediate ligand-dependent regulation of these receptors, and as such, LBDs have been the focus of intense structural studies. There are only a few orphan nuclear receptors for which the LBD structure remains unsolved. In the past two years, we have focused on structural characterization of two orphan receptors: constitutive androstane receptor (CAR) and steroidogenic factor-1 (SF-1). The CAR structure reveals a compact LBD fold containing a small pocket that is only half the size of the pocket in PXR, a receptor closely related to CAR. The constitutive activity of CAR appears to be mediated by a novel linker helix between the C-terminal AF-2 helix and helix 10. On the other hand, SF-1 is regarded as a ligand-independent receptor, but its LBD structure reveals the presence of a phospholipid ligand in a surprisingly large pocket; its size is more than twice that of the pocket in the mouse LRH-1, a closely related receptor. The bound phospholipid is readily exchanged and modulates SF-1 interactions with coactivators. Mutations designed to reduce the size of the SF-1 pocket or to disrupt hydrogen bonds formed with the phospholipid abolish SF-1/coactivator interactions and reduce SF-1 transcriptional activity. These findings establish that SF-1 is a ligand-dependent receptor and suggest an unexpected link between nuclear receptors and phospholipid signaling pathways.

The Met tyrosine kinase receptor

MET is a tyrosine kinase receptor that is activated by hepatocyte growth factor/scatter factor (HGF/SF). Aberrant activation of the Met receptor has been linked to the development and metastasis of many types of solid tumors and has been correlated with poor clinical prognosis. HGF/SF has a modular structure with an N-terminal domain, four kringle domains, and an inactive serine protease domain. The structure of the N-terminal domain with a single kringle domain (NK1) has been determined. Less is known about the structure of the Met extracellular domain. The molecular basis of the MET receptor–HGF/SF interaction and the activation of MET signaling by this interaction remains poorly understood. In collaboration with George Vande Woude and Ermanno Gherardi, we are developing this project to solve the crystal structure of the Met receptor/HGF complex.

G Protein–coupled receptors

G protein–coupled receptors (GPCRs) form the largest family of receptors in the human genome; they are receptors for diverse signals carried by photons, ions, small chemicals, peptides, and hormones. These receptors account for over 40% of drug targets, but the structure of these receptors remains a challenge because they are seven-transmembrane receptors. Currently, there is only one reported GPCR structure, for an inactive form of bovine rhodopsin. Many important questions regarding GPCR ligand binding and activation remain unanswered. From our standpoint, GPCRs are similar to nuclear hormone receptors with respect to regulation by protein-ligand and protein-protein interactions. Due to their importance, we have decided to take on studies of the structural basis of ligand binding in, and activation of, GPCRs.

External Collaborators

• Doug Engel, University of Michigan, Ann Arbor
• Ermanno Gherardi, University of Cambridge, UK
• Steve Kliewer, University of Texas Southwestern Medical Center, Dallas
• David Mangelsdorf, University of Texas Southwestern Medical Center, Dallas
• Donald MacDonnell, Duke University, Durham, North Carolina
• Stoney Simmons, National Institutes of Health, Bethesda, Maryland
• Scott Thacher, Orphagen Pharmaceuticals, San Diego, California
• Brad Thompson and Raj Kumar, University of Texas Medical Branch at Galveston
• Ming-Jer Tsai, Baylor College of Medicine, Houston, Texas