Submission Type
Poster
Abstract
The malate dehydrogenase (MDH) enzyme is found in a multitude of organisms functioning in the last step of the TCA cycle. MDH conducts the two-way reaction of malate and NAD+ to oxaloacetate and NADH (1). In humans, there are two major isoforms, cytosolic and mitochondrial. Deeper understanding of the MDH enzyme can expand treatments for parasitic diseases such as African Sleeping disease caused by T.brucei infection. Regulation of parasitic MDH gives the ability to limit the success of infection by inhibiting the protein’s ability to function (2). Since MDH can be controlled through competitive inhibition, drug discovery for T.brucei is the next step in MDH research (2). The objective is to characterize cMDH from T.brucei through protein expression and purification alongside kinetic assays using watermelon glyoxysomal MDH (wgMDH) as a control. Kinetic analysis was performed to obtain quantitative kinetic variables, and citrate was tested as a potential inhibitor for T.brucei cMDH. We were able to successfully express and optimize the purification of T.brucei cMDH, indicated by a pure and distinct band in the range of 34-37 kDa in SDS-PAGE. Bradford assay quantification of protein concentration for T.brucei cMDH was 7.5mg/ml and 9.7mg/ml for wgMDH. The kinetic data for cMDH and wgMDH for differing OAA concentrations revealed a Km of 760.3μM and 48.89μM and Vmax values of 203.3 μM/min and 80.72 μM/min, respectively. The kinetic data for differing NADH concentrations revealed a Km of 51.88uM and 248uM and Vmax values of 0.7594uM/min and 204uM/min for cMDH and wgMDH.
Recommended Citation
Sutherland, Grace; Jamburuthugoda, Varuni; and Bell, Ellis, "184 - Characterization of Cytosolic Malate Dehydrogenase of Trypanosoma brucei" (2025). GREAT Day Posters. 48.
https://knightscholar.geneseo.edu/great-day-symposium/great-day-2025/posters-2025/48
184 - Characterization of Cytosolic Malate Dehydrogenase of Trypanosoma brucei
The malate dehydrogenase (MDH) enzyme is found in a multitude of organisms functioning in the last step of the TCA cycle. MDH conducts the two-way reaction of malate and NAD+ to oxaloacetate and NADH (1). In humans, there are two major isoforms, cytosolic and mitochondrial. Deeper understanding of the MDH enzyme can expand treatments for parasitic diseases such as African Sleeping disease caused by T.brucei infection. Regulation of parasitic MDH gives the ability to limit the success of infection by inhibiting the protein’s ability to function (2). Since MDH can be controlled through competitive inhibition, drug discovery for T.brucei is the next step in MDH research (2). The objective is to characterize cMDH from T.brucei through protein expression and purification alongside kinetic assays using watermelon glyoxysomal MDH (wgMDH) as a control. Kinetic analysis was performed to obtain quantitative kinetic variables, and citrate was tested as a potential inhibitor for T.brucei cMDH. We were able to successfully express and optimize the purification of T.brucei cMDH, indicated by a pure and distinct band in the range of 34-37 kDa in SDS-PAGE. Bradford assay quantification of protein concentration for T.brucei cMDH was 7.5mg/ml and 9.7mg/ml for wgMDH. The kinetic data for cMDH and wgMDH for differing OAA concentrations revealed a Km of 760.3μM and 48.89μM and Vmax values of 203.3 μM/min and 80.72 μM/min, respectively. The kinetic data for differing NADH concentrations revealed a Km of 51.88uM and 248uM and Vmax values of 0.7594uM/min and 204uM/min for cMDH and wgMDH.
Comments
Sponsored by Varuni Jamburuthugoda