Lecture 8 Currency of metabolism ADP ATP NAD oxidized NADH reduced FAD FADH2 central metabolic pathways 1 glycolysis breakdown of glucose to pyruvate 2 pentose phosphate pathway 3 tricarboxylic acid cycle TCA Krebs Citrate cells generate energy impt for orgs that use O2 makes a lot of NADH FADH2 and little ATP 4 respiration conversion of organic molecs to CO2 O2 H2O 5 fermentation what happens in absence of O2 produce acids and alcohols chemoorganoheterotrophs catabolism of glucose 1 glycolysis 2 TCA 3 oxidative phosphorylation e transport chain and ATP synthase Glycolysis 10 steps uses 2 ATP make 2 NADH 4 ATP 2H2O net production gross amt invested amt net 2 ATP Gross 4 ATP mechanisms of active transport use ATP to transport molecs across membrane made to ADP 1 uniport 2 antiport 3 coupled transport 4 symport Substrate level phosphorylation occuring in glycolysis phosphates transferred from one substrate PEP to ADP to make ATP and pyruvic acid PEP ADP ATP pyruvic acid as ATP levels rise cells shut down phosphofructokinase enzyme when ADP is high and energy is low it activates enzyme pyruvate to acetyle coA pyruvic acid acetate CO2 acetate CoA acetyle CoA TCA Kreb s acetyl CoA in cycle of 8 parts make 1 ATP 3 NADH 2 CO2 all Xs 2 b c there are 2 pyruvates NADH converted into ATP through oxidative phosphorylation use electron transport chain and ATP Synthase Lecture 9 Glucose 2 pyruvate 2 ATP invested 4 ATP come out net 2 ATP 2 NAD 2 NADH Glycolysis Glucose 2 NAD 2 ATP 2 Pyruvate 4 ATP 2 NADH Substrate level phosphorylation 2 ADP Pi 2 ATP Oxidative phosphorylation 2 NADH 2 ATP 8 ATP Citric acid cycle pyruvate 4 NAD FAD GDP 3 CO2 4 NADH FADH GTP substrate leve phosphorylation 1 GTP 1 ADP 1 ATP 1 GDP oxidative phosphorylation 4 NADH 12 ATP 15 ATP x 2 2 pyruvates 1 FADH 2 ATP Glycolysis TCA 38 ATP Aerobic respiration breakdown of nutrients to produce energy using e transport chain O2 as final e acceptor substrate fully oxidized anaerobic respiration breakdown of nutrients to produce energy using e transport chain SO4 NO3 or others as final e acceptor substrate fully oxidized fermentation breakdown of nutrients to produce energy and recycles NAD use organic molecs as final e acceptor substrate partially oxidized regenerate NAD no additional ATP generated net gain from fermentation of glucose 2 ATP ATP yield Aerobic highest yield anaerobic fermentation lowest yield Protein protease amino acids hydrolysis Amino acid deaminated through hydrolysis and sent to Kreb s cycle if needed for energy Glucose NAD Pyruvic acid NADH Organisms NADH CO2 propionic acid lactic acid ethanol acetone isopropanol acetic acid NAD fermentation products Microbial replication 1 binary fission 2 budding most common for bacteria and archaea evenly split 1 DNA strand to mother and 1 DNA to daughter identical to parent cell common in yeast tiny object on mother cell separates and then grows to same size as mother 3 filamentous growth fungal growth grow in particular direction replication occurs on terminals grows in a line branches 30 min to double exponential growth chromosomes replicate septum forms septum completed splits generation time doubling time 1 2 4 8 16 32 etc between cell divisions synthesis of DNA ribosomes cell wall cell membrane cytoplasmic and outer proteins metabolism transport need twice as many of all of these phases of growth for batch culture 1 lag phase synthesis adaptation to environment sitting not many nutrients available b c used up immediate ones sensing environment and adjusting to it making DNA and energy 2 exponential phase log phase rapid increase in cell numbers after adaptation 3 stationary phase no net increase in numbers lack of available nutrients prohibits growth cell division rate death rate using dead cells around them as nutrients 4 death phase decrease in viable cell numbers death doubling senescence toxins that inhibit growth build up Lecture 10 Viable count Effect of temperature grow best at optimal temperature less more than optimal temp growth slows down Direct cell counts not all cells alive not all living cells will grow on growth media need high conc of organisms to get a count specialized chamber w grid etched count microorganisms use equation to convert square to cells microorg cells dead in gram stain Viable cell counts time consuming not all bacteria will grow need idea of of cells present to det the proper plating scheme use dilution spread plate Measure biomass need to know relationship b w bacterial size and cell count doesn t give actual of cells gives read out and have to convert media used can effect measurements need high concentration of cells take OD reading draw line down to viable count reading use graph to convert Measure cell products Toxic oxygen molecules 1 super oxide O2 2 peroxide H2O2 3 hydroxyl radical OH aerobes and O2 tolerant cells must have enzymes to get rid of these radicals superoxide dismutase and catalase superoxide H superoxide dismutase O2 H2O2 H2O2 catalase O2 H2O Physical methods of microbial growth control heat denatures proteins loses functions cells die can t make energy disrupts H bonds Oven dry heat req high temp long time Autoclave sterilization high pressure and steam 121 C help kill endospores use to sterilize culture media reusable medical instruments Pasteurization NOT sterilization used w heat sensitive liquids milk juice beer wine kills most organisms conditions designed to kill pathogens likely to be in sample filtration sample in filter in vacuum purity of sample depends on pore size of filter