Last Friday, students from all over the campus gathered at Weikamp in the morning and early afternoon to present research and creative works from the past year. As usual, our department was well-represented; this year our students contributed two talks and three posters. Pictured above-left is biochemistry major Maggie Fink with her mother on her right arm, while pictured above-right we have fellow biochemistry major Michele Costantino. Further down we see biology major Keon Jones shown with Associate Vice Chancellor (former organic chemistry professor) Doug McMillen. Keon presented his poster earlier this month at MoLSMAP in Missouri (click here for abstract). Jones and Costantino tied for the best poster presentation from the crowd of science and non-science posters.
And finally we find chemistry majors David Aupperle and Abigail Praklet - who both gave talks - pictured with biochemistry professor Shahir Rizk (who moderated the science session). They also traveled out of state in early April to present their work, but they went to NCUR in Oklahoma (click here for abstracts). Praklet's efforts won her the award for best talk in the science session which was an excellent way to cap her summer-fall-spring research adventure with Muna.
But all of our student researchers - whether they presented at the conference or not - deserve acknowledgement for their research efforts and have gained a valuable experience that other students may have yet to claim. If you are interested in research, please don't hesitate to ask about it. You can get the student point of view from fellow classmates - or visit one of your favorite professors to see what is happening outside of class.
Self Assembly of Novel Proteins Using Maltose Binding Protein and Engineered fABs
Maggie Fink, Professor Shahir Rizk
Many proteins undergo a large conformational change upon binding to a ligand. This conformational shift can expose protein surfaces previously shielded in the unbound state. A class of proteins that exhibits this conformational change is the bacterial periplasmic binding proteins. Each member of this family of proteins binds to a specific ligand, resulting in a shift from an open (unbound) to a closed (bound) conformation. Maltose binding protein undergoes this conformational change in response to maltose binding and transitions from an open to a closed form. To characterize the thermodynamics of these transitions and lock MBP into specific conformations, synthetic antibodies were designed to bind to MBP and screened in different maltose conditions to target both open and closed conformations. Three of these antibodies were isolated, one of which was found to bind to the open form of MBP endosterically (7O) and the other two, D1 and A1, bind to the closed form allosterically and peristerically. The aim of my project has been to construct a novel protein fusion where MBP and one of antibodies identified in the previous research are fused together. In this way the engineering proteins will potentially self-assemble into a nanostructure in the presence or absence of maltose, depending on the specificity of the antibody to the conformation of MBP. Once each of the three fusion proteins have been successfully cloned, expressed and purified, testing will be carried out to determine if a higher order complex can form in response to ligand addition or removal as a trigger.
The Effects of Metal Cofactors on Adenosine Deaminase Activity
Michele Costantino, Maggie Fink, Sandy Ho, Professor Shahir Rizk
Adenosine deaminase (ADA) is a zinc-dependent enzyme that converts adenosine to inosine as part of the purine degradation pathway. In humans, mutations in ADA are associated with severe combined immunodeficiency (SCID), an often fatal syndrome, due to accumulation of adenosine. This leads to B- and T-cell death, compromising the ability of the patient to fight infections. Previous work has shown that enzymatic activity depends on zinc concentration; however, at greater than 1:1 ratio, Zn can act as a negative allosteric effector of ADA activity. Other divalent cations — such as Co2+, Cu2+, Mn2+, and Cd2+ — can act as competitive or non-competitive inhibitors of the purified human enzyme. Additional studies indicate that Hg2+ inhibits crude ADA extracts from zebrafish at high concentrations. To characterize human ADA, kinetic studies with purified enzyme were conducted using direct colorimetric assays. Under optimal conditions, the rate of reaction was used to determine the KM and kcat of the uninhibited enzyme. The KM and kcat values were also obtained in the presence of coformycin, a known competitive inhibitor. The activity of ADA was measured in the presence of various divalent cations and Hg2+ was found to have the most profound negative effect on ADA activity. The inhibition of ADA by Hg2+ was found to be concentration dependent. This is the first study where the effect of Hg2+ on purified human enzyme was determined.
