Enzymatic Characterization of Aldose Reductase and Its Inhibitors

Enzymatic Characterization of Aldose Reductase and Its Inhibitors

Bibliographic Details
Main Author: Zivkovic, DaVena
Language:English
Published: Youngstown State University / OhioLINK 2016
Subjects:
Biochemistry
Analytical Chemistry
Aldose Reductase
Enzyme Kinetics
Thermogravimetric Analysis
Online Access:http://rave.ohiolink.edu/etdc/view?acc_num=ysu1472069987
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spelling ndltd-OhioLink-oai-etd.ohiolink.edu-ysu14720699872021-09-02T05:10:38Z Enzymatic Characterization of Aldose Reductase and Its Inhibitors Zivkovic, DaVena Biochemistry Analytical Chemistry Aldose Reductase Enzyme Kinetics Thermogravimetric Analysis Human aldose reductase (hAR) is a NADPH-dependent oxidoreductase that reduces aldehydes to their corresponding alcohols with the oxidation of NADPH. It has been shown that two forms of this enzyme can occur. When Cysteine-298 in the active site is mutated, different kinetic variables are experienced that correlate to the form of enzyme being obtained. Mutant C298A kinetic studies showed that the Michaelis-Menten constant, <i>K</i><sub>m</sub>, had an 18-fold increase, the turnover number, kcat, decreased about 83%, and the catalytic efficency, <i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub>, decreased about 100% for the mutant enzyme compared to the wildtype for the forward reaction using DL-glyceraldehyde as substrate and NADPH as cofactor. Mutant C298A kinetic studies showed that the Michaelis-Menten constant, <i>K</i><sub>m</sub>, had an 4-fold decrease, the turnover number, kcat, decreased about 60%, and the catalytic efficency, <i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub>, increased about 39% for the mutant enzyme compared to the wildtype for the reverse reaction using benzyl alcohol as substrate and NADP+ as cofactor. The mutation proved beneficial for the enzyme, shown to have been the targeted native form of the enzyme.Thermal analysis of clofibric acid samples in sodium chloride mixtures of 10% (w/w) ratios were investigated by thermogravimetric analysis and mass spectroscopy. Kinetic parameters like activation energy <i>E<sub>a</sub></i>, pre-exponential factor <i>A</i>, and order of reaction <i>n</i> were calculated for each sample's decomposition. Trends within the kinetic parameters occurred with increase of clofibric acid percentage. Mass spectroscopy data shows a detection of CO<sub>2</sub> with decomposition of clofibric acid samples. This data can provide insight on the possible mechanisms in clofibric acid decomposition. 2016-08-25 English text Youngstown State University / OhioLINK http://rave.ohiolink.edu/etdc/view?acc_num=ysu1472069987 http://rave.ohiolink.edu/etdc/view?acc_num=ysu1472069987 unrestricted This thesis or dissertation is protected by copyright: all rights reserved. It may not be copied or redistributed beyond the terms of applicable copyright laws.
collection NDLTD
language English
sources NDLTD
topic Biochemistry
Analytical Chemistry
Aldose Reductase
Enzyme Kinetics
Thermogravimetric Analysis
spellingShingle Biochemistry
Analytical Chemistry
Aldose Reductase
Enzyme Kinetics
Thermogravimetric Analysis
Zivkovic, DaVena
Enzymatic Characterization of Aldose Reductase and Its Inhibitors
author Zivkovic, DaVena
author_facet Zivkovic, DaVena
author_sort Zivkovic, DaVena
title Enzymatic Characterization of Aldose Reductase and Its Inhibitors
title_short Enzymatic Characterization of Aldose Reductase and Its Inhibitors
title_full Enzymatic Characterization of Aldose Reductase and Its Inhibitors
title_fullStr Enzymatic Characterization of Aldose Reductase and Its Inhibitors
title_full_unstemmed Enzymatic Characterization of Aldose Reductase and Its Inhibitors
title_sort enzymatic characterization of aldose reductase and its inhibitors
publisher Youngstown State University / OhioLINK
publishDate 2016
url http://rave.ohiolink.edu/etdc/view?acc_num=ysu1472069987
work_keys_str_mv AT zivkovicdavena enzymaticcharacterizationofaldosereductaseanditsinhibitors
_version_ 1719474166927720448

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