Respiration in Yeast and Mammalian Cells Respiration in Yeast and Mammalian Cells

C 6 H 12 O 6 + 6O 2 +6H 2 O6CO 2 +12H 2 O 36 ATP/glucose molecule Respiration: The cellular process by which glucose is oxidized to carbon dioxide and water. An inorganic molecule, often oxygen, serves as the final electron acceptor. The formation of ATP from this process can subsequently be used as the cell’s main source of energy 3 Step Process: 1-Glycolysis 2-TCA cycle 3-Electron transport chain Glucose

Glycolysis

Fermentation:

F 0 F 1 ATP synthase: Matrix Inner membrane Inner membrane space ADP + PiATP H+

Use mitochondria isolated from rat liver cells Day 2: Day 1: Use cultures of Sacchromyces cerevisiae

Glucose Pyruvate Acetyl CoA AcetaldehydeEthanol Electron Transport Glucose and Ethanol Utilization in Yeast NEM Azide

Experiment 1: Kinetics of respiration in fresh yeast Experiment 2: Kinetics of respiration in starved yeast Experiment 3: Effects of NaN 3 on respiration in starved yeast Experiment 4: Effects of NEM on respiration in starved yeast Day 1

Dissolved oxygen combines with platinum electrons + protons to form water Electrons move away from the silver electrode; positively charged silver ions combine with chloride ions to form silver chloride The remaining potassium ions combines with the hydroxyl ions from water The more oxygen there is, the faster the electrons move away from the silver electrode and the more current that is produced Its very important to keep the stirrer moving! Teflon membrane Sample chamber KCl Electrode Platinum (-) Silver (+) The Clark Electrode:

P/O ratio=ATP synthesized/O 2 utilized A B C D E F +medium +mitos +substrate +ADP Fraction of O 2 used CaCl time in minutes >3 indicates tightly coupled mitochondria Day 2: You will be testing various combinations of substrates, stimulators, inhibitors, and uncouplers on the rate of respiration

Antimycin A Glutamate,malate-complex I Succinate-complex II Ascorbate + TMPD-complex IV Azide OligomycinRotenone

Many of the chemicals you will be using are toxic Be Careful!!!!