The Consequences Of Inhibiting The Structure Of HREO ( TEO )

Succinate dehydrogenase is an enzyme found in the mitochondrial inner membrane. The enzyme catalyzes the reaction of oxidizing its substrate, succinate, into fumarate via the removal of hydrogen ions from succinate. This oxidation is vital in the Krebs cycle.

Since we have already known the amino sequence of the protein in previous step, we can narrow down the targeting ubiquitin ligase based on existing research data such as papers, NCBI data.

One of the best-studied peroxidases is horseradish peroxidase (HRP), which has a heme-iron co-factor. In most heme-peroxidases the iron atom in the active center undergoes a reversible change of its oxidation state. The reaction proceeds in three distinct steps. In first step, the resting state high-spin Fe(III) is present, which is oxidized by hydrogen peroxide to form an unstable intermediate called compound I (Co-I) with Fe(IV), releasing water in the process. Compound I is not a classical enzyme–substrate complex, but rather a reactive intermediate with a higher formal oxidation state (­5 compared with ­3 for the resting enzyme). Thus, compound I is capable of oxidizing a range of reducing substrates. This reactive intermediate oxidizes

In the respiration phase, citrate helps increase the intake of oxygen while slowing down the releasing of carbon dioxide as shown in Table I. Glycogen, hexosediphosphate, alpha-glycerophosphate increases the effect of the respiration phase helping support that carbohydrates are the substrates that are catalyzed by the citrate in the pigeon muscles. Next, the researchers examined the rate of disappearance of citric acid and found that the poisons arsenite or malonate make the citric acid in the muscle disappear in large groups if oxygen is present, but they do not affect the breakdown of citric acid. Table III shows the effect that arsenite has on the citric acid of the pigeon muscle. Researchers then looked at the conversion of citric acid to alpha-ketogluteric acid by using methylene blue to oxidize citric acid because neither poisons could fully oxidize citric acid (Table IV). The oxidation of the citric acid (malonate), is then checked by being placed in succinic acid, which identified succinic and alpha- ketogluteric acid by measuring the pressure of the fluid in a column or manometrically. To see if the synthesis of citric acid is true, the researchers must find out if citric acid can be regenerated through one of the products made through oxidation. The pigeon muscle was able to generate citric acid in the presence of oxaloacetic acid; however, the other intermediates including pyruvic acid

Urate hydroperoxide is formed when urate is oxidized to a radical that subsequently adds to superoxide. It is produced by activated white blood cells called neutrophils and by enzymes that include xanthine oxidase and peroxidase. Previous studies characterized the formation of urate hydroperoxide and its oxidation of small biomolecules (1-3). My work involved optimizing urate hydroperoxide production by xanthine oxidase and lactoperoxidase, and exploring how it reacts with thiols and thiol-dependent enzyme glyceraldehyde 3-phosphate dehydrogenase (GAPDH).

Enzymes are catalysts that lower the activation energy required to start a biological reaction and affects the rate of the rection. The enzyme can become denatured when its surroundings change in temperature or pH. Molecules in the environment could also affect the enzymatic activity. Inorganic substances known as cofactors and organic molecules known as coenzymes can enhance or inhibit the enzymes activity. The cofactors or coenzymes can act to activate or inhibit the enzymatic activity. Peroxidase is a catalyst that catalyze substrate oxidation when a peroxide is present. The indicator guaiacol is easily oxidized by peroxidases. Guaiacol is used to indicate if the peroxidase enzyme is present in a solution. Turnip peroxidase breaks down hydrogen peroxide into water and

Oxidants will affect mitochondrial redox status and may cause extensive oxidative damage to proteins(Kalyanaraman et al .,2002). Furthermore, the mitochondrial transition pore can open releasing cytochrome c and possibly affecting mitochondrial function. Cytochrome c can activate cytosolic caspases to induce apoptosis. Doxorubicin may induce anti-apoptotic (Bcl-2, Bcl-XL) and pro-apoptotic (Bax, Bad) proteins. The Bcl-2 family of proteins regulates the release of cytochrome c . These include Bax, Bad, and Bid, which are pro-apoptotic proteins that favor cytochrome c release and Bcl-2 and Bcl-XL which are anti-apoptotic proteins that inhibit cytochrome c release . The mechanisms of how Bcl-2 family proteins regulate release of cytochrome c and apoptogenic factors are under investigation.

Fig. 6 A simplified illustration of electron transport chain pathways highlighting the effect of tamoxifen. Tamoxifen was showed to be a strong uncoupling agent and an inhibitor of electron transport chain function. More specifically, it inhibits the activities of complex II+III (IC50 =15µM) and complex V (IC50 =8.1µM)[6]. Adapted from publication of Nadanaciva et al. [6].

Reduction of an electron from ring C in ANT causes to formation of free radical semiquinone. This radical is partly stable in anoxic environment but under normoxic condition, its unpaired electron is given to oxygen and superoxide radicals are formed. Appropriate flavoproteins such as complex I catalyses reduced semiquinone radicals by accepting electrons from NADH or NADPH and delivering them to ANT. This sequence of reactions are known as “redox cycling” that can be very deterious (dangerous) because low amount of ANT is adequate for formation of many superoxide radicals (67). This radical damage triggers production of highly toxic aldehydes such as malondialdehyde (MAD). These aldehydes can diffuse easily in the cell and even from cell membrane

First classical methods are described. The CHARMM force field, which we have used in this thesis is also briefly described. Then the quantum mechanical methods such as ab initio and semiempirical methods are briefly described. The SCC-DFTB method is described more elaborately since we have used SCC-DFTB as the quantum mechanical method to study the reaction mechanism in this thesis. Then QM/MM methods are described with available methods to study enzymatic reaction mechanisms.

For cellular respiration the mitochondria were expected to respire, while the inclusion of succinate would be necessary for the oxidation of DPIP (Upadhyaya, 2017). These hypotheses were proven to be correct. In the fermentation lab the metabolic rate was recorded by measuring the amount of CO2 released, which differentiated among the different carbohydrates and temperatures which was attributed to the one being a monosaccharide (glucose), a disaccharide (sucrose), and a polysaccharide in the form saturated starch. The simple sugars had a faster metabolic rate because their bonds were more readily broken down. In the cellular respiration lab it was shown that mitochondria respire regardless of the addition of succinate, however, when succinate is added it would increase the rate of respiration.

The protein usually targets old, dysfunctional proteins from the cell cycle, which makes sense for controlled cell division. The ubiquitination mechanism was originally obscure, but made more clear in the mid-1980’s by Aaron Ciechhanover, Avram Hershko, and Irwin Rose, which helped them win the Nobel Prize for Chemistry in 2004 (Nobel Prize website). Initially, the process was looked at as proteolytic system utilizing ATP. Many ATP-dependent proteolysis factor 1 (APF-1) molecules linked with a substrates that were later found to be deteriorated with free APF-1 in the cellular extract (Ciechhanover, 2012). APF-1 was called ubiquitin and it uses its C-terminal glycine to conjugate to the substrate’s

They are uncoupling proteins that work together to control mitochondrial ROS formation. They work to move protons from the MIM space into the matrix by using the electrochemical gradient present. For UCP2 and UCP3 the process of reversible glutathionylation of the 2 proteins regulates the activation (UCP2) and deactivation (UCP3) of the leak.

There is a class of related enzymes known as Phenolases. These enzymes catalyze the oxidation of compunds that have hydroxyl groupds attatched to a benzene ring, recongized as phenols. The enzyme is shown to be nonspecific and capable of oxidizing Substrates with acidic side chain are oxidized less effectively having the possibility of denaturing the enzyme. The substrate within the enzyme is oxidized, or combine or become combined chemically with

A catalase is an enzyme that protects cells from oxidative damage. It does so by breaking down hydrogen peroxide, which the catalase generates during cell metabolism. Catalase deactivates millions of hydrogen peroxide molecules per second. First, the catalase breaks down hydrogen peroxide by removing and binding one oxygen atom and next releasing the rest of the hydrogen peroxide molecule as water. Then, the catalase breaks down a second hydrogen peroxide molecule by releasing oxygen gas and water. The optimum temperature- the temperature at which the enzyme is the most active at- is 40°C. Also, the optimum pH level for the catalase is pH 7.