Research Interests

• The Dynein Motor Protein Complex
• Protein Structure and Dynamics
• Protein Folding
• Protein-Protein Interactions

Cytoplasmic dynein is a biological system composed of a number of proteins acting together in a very precise way. To understand this system, we use a 'ground up' approach, whereby the structure and dynamics of the individual proteins are examined in detail, then the interactions between subunits is characterized. Our efforts are guided by the fact that our proteins are derived from a living system, where their structure has been optimized to perform a specific function. Our overriding goal is to understand relationships between structure and function. We have the ability, through a collaborator at the University of Minnesota, to incorporate modified proteins into a living organism (Drosophila) to test hypotheses about structure/function relationships.

Dynein is a motor protein, with the remarkable ability to convert the energy stored in ATP into a mechanical force capable of moving material through the cell along filaments called microtubules. We are particularly interested in cytoplasmic dynein, a motor protein which is involved in the placement of chromosomes along the spindle during cell division. Other processes which require active transport within the cell are also known to use the dynein motor.

Cytoplasmic dynein is a large complex composed of two heavy chains and a number of smaller light and intermediate chains. The figure shown to the right is a schematic diagram based on electron micrographs of the dynein particle. The heavy chains contain the ATP hydrolysis sites, and 'walk' along the microtubules. The role of several of the smaller subunits has so far only been surmised from their location within the cell. They are believed to be responsible for recognition and attachment of the dynein complex to a specific cargo at the proper time. We have focused our efforts on a few subunits which are known to be essential to several organisms - If the protein is not expressed, the organism has limited viability.

Heteronuclear NMR has become one of the primary tools in structural biochemistry, and our group is one of the heaviest users of the departmental NMR facility, as well as other state-of-the-art facilities. NMR is used to determine the three-dimensional structure of proteins, which is the basis for prediction of its properties. With detailed structural information we can, for example, confidently predict the sites where a protein will interact with other proteins and small molecules, and we can make informed inferences as to the forces which stabilize its unique functional structure. Like NMR, recent developments have made mass spectrometry an essential element of protein research. We use an electrospray mass spectrometer not only to verify the composition of proteins we have expressed and purified, but also to gain valuable structural information using hydrogen isotope exchange techniques.

Since fluorescence spectroscopy is sensitive to conformational changes of proteins, it is extremely valuable in tracking changes in protein structure as a function of temperature and solvent conditions. For example, we have recently used fluorescence to support our hypothesis that the LC8 subunit has a different quaternary structure in different environments within the cell.

In the end, we hope that our work on dynein will lead to new methods for controlling certain diseases. (The cancer drug Taxol is effective in preventing spindle formation in rapidly dividing cells; a complementary therapy would prevent transport of chromosomes by dynein along the spindle.) Perhaps just as important, however, will be advances in our fundamental understanding of protein-protein interactions and the assembly of biologically important protein complexes.