Microbial Metabolism

What is metabolism? Sum total of ALL chemical reactions in a living organism Metabolism is about the energy balance in cells, production and use CATABOLIC reactions release energy ANABOLIC reactions require energy These processes are COUPLED

Coupled Reactions

Enzymes Enzymes are biological catalysts Enzymes lower the activation energy required for biological reactions to occur Enzymes are: –Proteins –Specific for the reactions they catalyze –Not used up in the reactions –Speed up reactions in living system –Generally end with -ase

Energy Requirements

Enzymes Enzymes are proteins Many enzymes are part protein and part something else Protein part is called the APOENZYME Non-protein part is called a COFACTOR Cofactors are often metal ions If the cofactor is an organic molecule it is called a COENZYME Together the apoenzyme and cofactor form the HOLOENZYME

Holoenzyme

Coenzymes Coenzymes are organic molecules Often derived from vitamins Some coenzymes act as electron carriers in metabolic reactions Two important coenzymes in metabolism are: –NAD+ (nicotinamide adenine dinucleotide) –FAD (flavin adenine dinucleotide) –Both are electron carriers

Enzyme action Enzymes have a specific region on the surface where reactions occur ACTIVE SITE Enzyme-substrate complex forms at the active site Reaction occurs Products are released

Enzyme action

Factors that influence enzyme activity Temperature pH Substrate concentration Inhibitors

Temperature, pH, Substrate

Enzyme Inhibition Competitive inhibition Inhibitor “looks like” the natural substrate and fills the active site preventing the enzymatic reaction Noncompetitive inhibition Inhibitor binds to another site on the enzyme ALLOSTERIC SITE Causes a change in the active site and loss of activity

Inhibition

Competitive Inhibition Sildenifil citrateCyclic GMP

How to make ATP First you need ADP Next some phosphate Add some energy → ATP ADP + P +energy → ATP But where does the required energy come from? That’s the next slide…….

Where’s the energy? Adding the phosphate to ADP to make ATP is called PHOSPHORYLATION Cells do this in two ways: 1. Substrate level phosphorylation - the phosphate is transferred directly to the ADP to make ATP 2. Oxidative phosphorylation – electron carriers such as NAD+ transfer electrons to a series of carries in the electron transport chain where energy is released and transferred to ADP to make ATP

Oxidation and Reduction Part of the energy production mechanism in cells Oxidation = loss of electrons Reduction = gain of electrons These reactions are always coupled in cells Oxidation reactions produce energy

Redox reactions

Electron Carriers of Oxidative Phosphorylation NAD+ –NAD+ gains 1electron and 1H+ NADH FAD –FAD gains two H atoms FADH2 –The electrons will eventually be passed through the electron transport system to generate energy

Carbohydrate catabolism Cells break down glucose to make energy Two processes are involved: 1. Cellular respiration 2. Fermentation Glycolysis Krebs cycle Electron transport chain

Respiration and Fermentation

Glycolysis Means glucose burning Occurs in either the presence or absence of oxygen Yields relatively little ATP Most living cells use this process

Glycolysis

Out come of glycolysis Start with 1 molecule of glucose (6C) The products are: –NET 2 molecules of ATP –2 molecules of NADH –2 molecules of pyruvic acid (3C) Organisms that are aerobic or facultative proceed into the Krebs cycle and ETS Anaerobes proceed to fermentation

Krebs Cycle

How are Glycolysis and the Krebs Cycle connected? Pyruvic acid is the END PRODUCT of glycolysis Pyruvic acid itself DOES NOT directly enter the cycle It is first converted to acetyl CoA, and this compound enters the Krebs cycle Pyruvic acid → acetyl CoA yeilds: –NADH x 2 = 2 NADH –CO2 x 2 = 2 CO2

Krebs Cycle Aerobic process Has the potential to generate the most energy for the cell The products of this cycle are: –3 molecules of NADH x 2 = 6 NADH→ ETS –1 molecule of FADH2 x 2 = 2 FADH2 →ETS –1 molecule of ATP x 2 = 2 ATP Why x 2? Because glycolysis generates 2 molecules of pyruvic acid as an end product so the cycle must “go around” twice to metabolize all the pyruvic acid!!

Electron Transport Chain of carrier molecules that receive electrons from NADH. The flow of electrons from carrier to carrier is exergonic. The energy release is used to pump H+ from NADH across a membrane to form a concentration of H+ on one side of the membrane. The flow of H+ back across the membrane is also exergonic and the energy released is used to convert ADP to ATP – PROTON MOTIVE FORCE! ATP is made via CHEMIOSMOSIS by the enzyme ATP SYNTHASE Final electron acceptor is oxygen ETS occurs only in the presence of oxygen The reaction to make ATP is called OXIDATIVE PHOSPHORYLATION. The source of electrons are the NADH and FADH2 produced in earlier steps of glycolysis and the Krebs cycle

Electron Transport System

ATP Production by ETS For every 1 NADH that goes into ETS you get 3 ATP For every 1 FADH2 that goes into ETS you get 2 ATP How many NADH and FADH2 are generated in ALL parts of AEROBIC respiration? ANSWER 10 NADH and 2 FADH2

Total Energy Production 1 molecule of glucose gives the following: 10 NADH → ETS = 30 ATP 2 FADH2 → ETS = 4 ATP 2 ATP directly from glycolysis (substrate level phosphorylation) 2 ATP directly from Krebs (substrate level phosphprylation) TOTAL ATP from 1 glucose = 38

Summary of aerobic respiration

What about the anaerobes and energy? If you are an anaerobe you can do only glycolysis and get how munch total ATP? Right, just 2 ATP Remember no ETS for these microbes! But want about the end product pyruvic acid, what happens to it, it can’t just continue to build up? Answer, these bacteria use the pyruvic acid in the process of FERMENTATION

Fermentation Occurs in the absence of oxygen Produces small amounts of ATP Organic molecule, not oxygen, is the final electron acceptor End products are alcohol or various acids Anaerobes and facultative anaerobes

Let’s make wine! What do you need? Grape juice, that’s the source of sugar Yeast, these cells will make the alcohol Mix together and put in a bottle that can be sealed What’s happening in the bottle? The answer is fermentation – no oxygen, you sealed the bottle, the yeast break down the sugars in the grape juice, make pyruvic acid, then alcohol Wait a few weeks, open the bottle, then PARTY!

What wine? The party was no fun, there was no wine What happened, yeast make alcohol don’t they? Right, they do, but only if NO oxygen is present You didn’t seal the bottle well, air got in and the yeast did this: –Grape juice→glycolysis→Krebs→ETS –NO alcohol!

Fermentation

End Products

Catabolism

Nutritional Classification

Energy comparisons Aerobes and facultative anaerobes growing the presence of oxygen produce 38 ATP from one molecule of glucose Anaerobes produce 2 ATP from one molecule of glucose So the energy bottom line is that growth in the presence of oxygen is the way to go and grow!