Associate Professor of Practice
Degrees and Appointments
- Ph.D. Chem. Eng., University of California, 2000
- B.E. Chem. Eng. with Distinction, University of Delaware
We have continued and plan to expand the guided inquiry research component of our one semester Biochemistry Laboratory Course (BIOC463A). Since 2009, the second portion of the semester involved a Special Research Project. Initially our studies involved examination of the structural importance and roles of two disulfide bonds in alkaline phosphatase, a periplasmic enzyme from E. coli, using the chemical reducing agents beta-mercaptoethanol, dithiothreitol, and triscarboxyethylphosphine. These early studies included a variety of spectroscopic methods including activity assays, fluorescence spectroscopy, and circular dichroism (CD) spectroscopy.
Our attempts at incorporating real research into our course has been highlighted in several posters presented at the American Society for Biochemistry and Molecular Biology (ASBMB) annual meetings as well as two invited talks describing our transformation process.
In 2013 Dr. James Hazzard began a two year plan to develop the current protein expression system, which in fall 2014 was realized with the designed pETHSUL-AP vector that has successfully incorporated into Origami cells for the production of disulfide bond containing proteins. In the spring 2015 semester we designed our first set of site-directed mutants of AP using PCR mutagenesis techniques, which is being done by each section of BIOC463A. Our most recent work has helped us quantitatively assess the activity of our wild type and mutant enzymes from the Origami cells compared to commercially available sources. Recently, we have found a strong correlation between the dimeric state of the enzyme, it's overall activity and the presence of one of the disulfide bonds.
Since 2018, we have started working on oligomerizing enzymes and their regulation. A new developing area in biology is the study of how the oligomeric state of a protein, particularly in either filamented or phase separated states, has dramatic effects on activity. One might think that many filamented or "aggregated" enzymes are merely a storage depot for unneeded reactions. However, about half the filaments studied structurally are more active in the filamented state. We are actively engaged in studying the filamenting of the human phosphofructokinase isozymes and how this regulates their activity in vivo. We approach the problem with classical protein production and biophysics. The addition of coupled reaction assays to the lab class has helped broaden
Finally, the lab has a long history with analytical ultracentrifugation (AUC) as well as vesicles, liposomes, and eventually, exosomes. We have begun using the AUC and other traditional characterization techniques to survey the distribution and contents of these cellular microenvironments.