Tracy Handel

Paying the bills and cracking the whip

Professor
University of California, San Diego
Skaggs School of Pharmacy and Pharmaceutical Sciences
9500 Gilman Drive, MC 0684
La Jolla, CA  92093-0684

Phone:      858.822.6656
Lab:        858.822.6606
Fax:        858.822.5591
E-mail:     thandel@ucsd.edu

Samantha Allen

Structure/function studies of chemokine receptors

Structural and functional studies of chemokine receptors would clearly help our understanding of both the molecular details of chemokine-receptor signalling and the roles of chemokine receptors in disease. A number of high-resolution structures of chemokines have been solved and many functional studies have been undertaken, but much less is known about their receptors. This is despite the estimation that >60% of the drugs on the market are directly or indirectly targeted at GPCRs. This lack of knowledge regarding membrane proteins is not surprising, because of the challenges associated with their expression and functional reconstitution, but it is necessary to develop methods in order to improve our understanding of these proteins. To this end, I'm using insect and mammalian cell expression systems to develop ways to express, purify, and reconstitute chemokine receptors, for structural and functional studies.

Xiyun Zhang

Computational active site design

My research focuses on the computational design of enzyme catalysts for non-natural reactions of synthetic significance, such as a Diels-Alder reaction. The research is the combination of chemistry, biology, computer science, and engineering. We are competing with nature, which perfected natural enzymes by evolutionary pressure over millions of years. This is a daunting task that is now becoming feasible, as success were achieved recently in designing new protein scaffolds, biosensors and enzymes.

My research is based upon the EGAD library developed in our lab for rational protein design with the new combination of quantum mechanical (QM) calculations for enzyme active site design. The design approach involves first building the transition state (TS) surrounded by a unique catalytic site. The construction is based upon known catalytic mechanisms and QM calculations. Following "flexible active site grafting" will search the catalytic site against the Protein Data Bank for matched backbones that do not have clashes with the TS and catalytic site. Subsequent core-repacking and sequence optimizing with EGAD will generate a protein with properly oriented catalytic groups in the active site. Proficient QM/MM methods, where the bond making-and-breaking TS and the residues participating in catalysis are modeled with QM methods, and the remainder of the enzyme is treated less intricately using molecular mechanics (MM), will be employed to predict which of the designed proteins are likely to be the best catalysts. The final candidate proteins will then be synthesized and tested for folding and catalytic activity, and if successful, structurally characterized.

Arnab Chowdry

Computational design of ligand binding and active sites

Proteins are a versatile system for the design of molecular machines, both in-vivo and in-vitro . The creation of catalytic antibodies against biologically irrelevant transition state analogues to produce novel enzymes suggests that proteins are capable of much more than their biologically relevant roles. Certain protein scaffolds, such as the TIM barrels, are known to be used for many distinct enzymatic roles. I propose to harness the TIM barrel scaffold for the design of novel enzymes.

Andrew Douglas

Structure and binding of M11L

I am presently studying the viral inhibitor of apoptosis M11L. M11L is expressed by the Leporipox virus Myxoma, and is a potent inhibitor of apoptosis. M11L is known to directly bind to the "death" protein Bak, and to a lesser extent to Bax, and inhibit its function in killing cells. Based on secondary structure predictions and M11L's ability to bind bak, it is likely a member of the Bcl-2 family of proteins. It is interesting to note that M11L show no sequence similarity to any member of the Bcl-2 family. It is my interest to work on solving the structure of M11L, to determine if it is in fact a structural homolog of Bcl-2. In addition I would like to explore the binding interaction between M11L and the other Bcl-2 proteins in hopes of understanding the global function of M11L.

Melinda Hanes

Directed evolution of the BLIP/ß-lactamase system

Bacterial resistance to antibiotics is an increasing problem in the medical community for a number of reasons. An important example is the production of ß-lactamases which inactivate ß-lactam antibiotics, such as TEM-1, or SHV-1. Combining antibiotics with small molecule inhibitors of these enzymes sometimes effective, but inhibiter resistant ß-lactamases are evolving. ?-lactamase inhibitor protein (BLIP) binds many lactamases with affinities ranging from nanomolar (TEM1) to micromolar (SHV1). By using an irrational design approach, I hope to find a BLIP variant that binds tightly to and inhibits SHV-1. Specifically, the experimental approach will be phage display, a powerful directed evolution technique. The results will lead to understanding and characterizing the critical interactions in the BLIP/ß-lactamase system.

Andro Hsu

Alanine scanning of CTACK/CCL27

I am performing an alanine scan of the chemokine CTACK/CCL27, a ligand of CCR10, with the goal of identifying the receptor and glycosaminoglycan- (GAG-) binding epitopes. CCR10 is overexpressed in melanoma cells, and the action of CTACK may be responsible for the preferential metastasis of melanoma to tissues and organs in which CTACK is constitutively expressed. Ultimately, I hope to identify a mutant of CTACK that is an antagonist for CCR10, so that CTACK's role in the metastasis of melanoma can be demonstrated in an in vivo mouse model.

Ariane Jansma

NMR spectroscopy of membrane proteins

I spent three years working as an NMR spectroscopist at both DuPont Pharmaceuticals and GNF in San Diego, while finishing my MS in analytical chemistry. My goal in coming back to graduate school to pursue a PhD was to learn to apply NMR spectroscopy to biological systems, specifically membrane proteins, as well as expand my knowledge of techniques in both molecular and cellular biology. As a first year graduate student in the Handel Lab, I am interested in studying the structure and function of various chemokines and particularly their membrane bound receptors. For the moment, I am working in collaboration with Sam Allen to study the chemokine receptor D6 as well as the NMR analysis of the Tig2 protein. In addition, I am assisting Andro Hsu on the expression and NMR analysis of the chemokine MEK. My immediate goal is to use these collaborations to gain new skills sets and to begin development of my own project.

Kim Reynolds

Structural and computational studies of the BLIP/ß-lactamase system

Protein-protein interactions are critical for everything from inter- and intra-cellular communication and regulation events to cytoskeletal structure. Efforts to characterize the interactions stabilizing protein interfaces have revealed few general rules for creating a stable interface. Instead, it appears that stable, specific complexes may arise from a diverse array of structural interactions. One interesting and potentially informative system for studying protein-protein interactions is the interface formed between ß-lactamases and ß-lactamase inhibitor protein (BLIP). BLIP is capable of binding and inhibiting both TEM1 and SHV1 ß-lactamase. These two ß-lactamases mediate antibiotic resistance in gram negative bacteria. Despite the fact that TEM1 shares 66% sequence identity with SHV1, and their crystal structures overlay with an RMSD of 1.4 Å (0.65 Å at the interface residues), BLIP exhibits a 1,000 fold difference in binding affinity for the two. While BLIP binds TEM1 with nanomolar affinity, it binds SHV1 with only micromolar affinity.

I am interested in dissecting the interactions involved in binding affinity and specificity for these complexes through traditional methods such as X-ray crystallography and mutagenesis, as well as through the use of computational protein design. Using the EGAD algorithm for protein design, I hope to identify residues important for binding affinity in the two complexes, and design tighter-binding BLIP variants for both SHV1 and TEM1.