Rachel Kooistra Thesis Defense: Project Description
Title: Functional characterization of a novel thioredoxin-containing protein of the malaria parasite Plasmodium
A novel thioredoxin domain-containing protein of the malaria parasite Plasmodium was identified and found to be conserved among eukaryotes. This protein belongs to the phosducin-like family of proteins (PhLPs), and was therefore assigned the name PhLP1, since it is the first phosducin-like protein to be identified in Plasmodium. PhLPs have been found to have various roles in G-protein signaling, cell cycle progression, and protein folding. However, the biochemical mechanism by which PhLPs perform their function is unknown. Here is described the cloning and biochemical characterization of both PhLP1 and its human homolog TXNDC9. Both purified PhLP1 and TXNDC9 showed enzymatic activity in the insulin reduction assay and were also active in the thioredoxin-coupled reduction assay. Sequence alignment and homologous modeling of PhLP1 and TXNDC9 indicated a conserved, putative atypical active site in place of the typical catalytic CXXC motif found in classical redox active thioredoxins. Site-directed mutagenesis of the putative redox active cysteine (C106) in PhLP1 abolished the redox functions of the protein, confirming the role of C106 in the catalytic mechanisms of the protein. These results show for the first time that PhLPs are redox active enzymes that can be efficiently reduced by the thioredoxin system. These findings shed new light on the biochemical mechanism and biological function of these highly conserved proteins.
First and foremost, I would like to thank my thesis advisor, Dr. Stefan Kanzok. His door was always open for questions and concerns, and he always encouraged me to come up with my own solutions and hypotheses. I am especially grateful for his continuous support of my career goals, and for his willingness to listen to my complaints when nothing seemed to be working out in lab. In addition, a huge thank you to every member of the Kanzok lab (past and present), but especially to Robin David, Katie Fell, Kyle Haselton, Sana Hira, Mike Lamm, and Shannon McGuire. Thank you also to my committee members, Dr. Rodney Dale, Dr. Catherine Putonti, and Dr. Kim Williamson, for their invaluable advice. Finally, thank you to other contributors to this project, Dr. Kenneth Olsen, Dr. Andrew Blagborough, and Dr. Holly Goodson.
Rachel Kooistra was born January 26, 1989, in Rockford, Illinois. She graduated in 2011 from Loyola University Chicago with a Bachelor of Science degree in Biology. In August of 2011, she entered the Master of Science program at Loyola University Chicago. Rachel was awarded a Graduate Fellowship for 2011-2013.
Dr. Stefan Kanzok
Dr. Rodney Dale
Dr. Catherine Putonti
Dr. Kim Williamson