Date Thesis Awarded

5-2020

Access Type

Honors Thesis -- Access Restricted On-Campus Only

Degree Name

Bachelors of Science (BS)

Department

Chemistry

Advisor

Lisa Landino

Committee Members

Douglas Young

Paul Heideman

Abstract

Lactate dehydrogenase (LDH) is an enzyme that interconverts lactate and pyruvate making it a critical regulator of cellular metabolism. LDH is also involved in several other signaling pathways due to its ability to bind RNA and ssDNA. The dysregulation of cellular metabolism can be seen in several diseases including cancer. There are two major isozymes of LDH, LDH-A and LDH-B which are the focus of this study. Native gel electrophoresis and spectrophotometer based enzyme activity assays were used to investigate conformational changes in LDH isozymes. Cytochrome c (cyt c) is a protein involved in the electron transport chain and is critical for apoptosis. When LDH-A is exposed to cyt c the conformation of LDH-A changes, and there is a 17% increase in LDH-A activity. This activity increase is not seen in LDH-B. LDH-B did not change conformations due to any protein tested in this study suggesting a more rigid structure than LDH-A despite their similar amino acid sequences. The change in conformation of LDH-A is reduced after cyt c denaturation which suggests the secondary and tertiary structures of cyt c play a role in the interaction. When LDH-A and LDH-B were exposed to cofactors (NAD + /NADH), LDH-A increased in activity whereas LDH-B decreased in activity. These results demonstrate multiple ways in which the cellular environment, whether that be other proteins or cofactors can influence the conformation and activity of LDH. Gaining a better understanding of how critical regulatory proteins interact with their environment is important for the design of novel therapies.

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