Microbial Metabolism:
Catabolic and Anabolic Reactions
Metabolism: the sum of the chemical reactions in an organism.
Catabolism:
C6H12O2+ O2->CO2+H2O (exothermic, exergonic)
Anabolism:
energy+CO2+H2O->C6H12O6+O2 (Endothermic, Endergonic)
Provides energy and building blocks for anabolism. (makes smaller molecules)
Uses energy and building blocks to build large molecules.
Metabolic Pathway
a Sequence of enzymatically catalyzed chemical reactions in a cell
determined by enzymes
encoded by genes
Collision Theory:
Activation Energy:
Reaction Rate:
When atons, ions, and molecules collide.
disrupts electonic configurations
is the frequency of collisions with enough energy to bring about a reaction
The reaction rate can be increased by enzymes or by increasing temperature or pressure.
Enzyme Components:
Biological catalysis
- specific for a chemical ration, not used up
Apoenzyme: protein
Cofactor: nonprotein component
-coenzyme: organic cofactor
Holoenzyme: apoenzyme plus cofactor
Important Coenzymes
NAD+
NADP
FAD
Coenzyme A
Enzyme Classification
Oxidoreductase: oxidation-reduction reactions
Transferase: transfer functional groups
Hydrolase: Hydrolysis
Lyase: removal of atoms without hydrolysis
Isomerase: rearrangement of atoms
Ligase: joining of molecules;use ATP
Factors Influencing Enzyme Activity
Temperature
pH
Substrate concentration
Inhibitors
* Temperature and pH denature proteins
Enzyme Inhibitors: Feedback Inhibiton
...
Ribozymes
Cut and Splices RNA
Oxidation-Reduction Reactions
Oxidation: removal of elections
Reduction: gain of electrons
Redox reaction: an oxidation reaction paired with a reduction reaction
Associated with hydrogen atoms
Biological oxidations are often dehydrogenations
The Generation of ATP
ATP is generated by the phosphorylation of ADP
Substrate -Level Phosphorylation
Energy from the transfer of a high-energy PO4-, tp ADP generates ATP
Oxidative Phosphorylation
Energy released, generat ATP in the electon transport chain.
Carbohydrate Catabolism (learned in Bio 156)
The Breakdown of carbohydrates to release energy
- Glycolysis
-Krebs cycle
- Electron transport chain
Figure 5.11
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A summary of Respiration
Aerobic Respiration- electron acceptor
Anaerobic Respiration: NOT O2
- yeilds less energy then aerobic respiration
Anaerobic Respiration
Electron Acceptor Products
NO3- NO2-,N2+H2O
So4- H2S+H2O
CO32- CH4+H2o
Carbohydrate Catabolism
Pathway Eukaryote Prokaryote
Glycolysis Cytoplasm Cystoplasm
Intermediate Cytoplasm Cytoplasm
Krebs cycle Mitchondrial matrix Cytoplasm
ETC Mitochondrial inner membrane
Plasma membrane
The Electron Transport Chain
electrons are passed down the chain
energy released can be used to produce ATP by chemiosmosis
Fermentation
spoilage of food
produces alcoholic beverages
large scale microbial process occurring with or without air.
-releases energy from oxidation of organic molecules
-does not require oxygen
-Does not use the Krebs cycle or ETC
- Organic molecules as the final electron acceptor
*Alcohol fermantation: produces ethanol +CO2
* Lactic acid fermantation: produces lactic acid
Chemotrophs
Use enegery from chemicals
Chemoheterotroph
Chemotrophs
Energy is used in the Calvin-Benson cycle to fix CO2
Phototrophs
Use light energy
Photoautotrophs use energy in the Calvin-Benson cycle to fix CO2
Photoheterotrophs use energy
Figure 5.27 Requirements of ATP production
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Figure 5.28 A nutrional classification of Orgnaisms
* ALL ORGANISMS HAVE TO HAVE ENERGY
-CHEMICAL
-LIGHT
Metabolic Diversity among Organisms
...
Biological oxidation are also called ___ reactions.
dehydrogenations
At the completion of the Krebs cycle, the carbons from glucose are in ___ (name the molecules)
Carbon dioxide (co2)
At the completion of aerobic respiration,energy has been formed. The energy from the oxidation of glucose is sotred in ____?
ATP
_______ involves an inhibitor that fills the active site of an enzyme and competes with the normal substrate for the active site.
Competive inhibition
For every NADH that is oxidized via the electron transport chain,___ ATP are formed.
3
All of the following pairs are correctly matched Except
Oxidation: reaction where are gained
Competitive inhibition of enzyme action evolves
Competition with the substrate for binding at the active site.
During glycolysis, elctrons from the oxidation of glucose are transferred to
NAD+
Streptococcus (In italics) bacteria lack an electron transport chain. How many molecules of ATP can a Streptococcus (In italics) cell net from one molecule of glucose?
2
Unlike eukaryotes, in prokaryotes chemiosmosis
Occurs at the plasma membrane an not the mitochondria.
The chemical reactions involved in synthesizing proteins and cell wall peptidoglycan are examples of _____ reactions.
anabolic
Fopre a cell wall, which of the following compounds has the greatest amount of energy per molecule?
Acetyl CoA pyruvate
The complete oxidation of glucose typically involves which three stages?
Glycolysis,Krebs cycle, and the elctron transport chain.
Which of the following mechanisms does not generate ATP using an electron transport train?
Substrate-level phosphorylation
Which of the following are products of the light- dependent reactions, on which the light dependent reactions are dependent.
ATP and NADPH
Enzymes work most effectively at their optimal temperature and pH.
True
Feedback inhibition generally acts on the last enzyme in an anabolic pathway
False
Glycoysis produces ATP through substrate phosphorylation
True
The synthesis of sugars by using carbon atoms from Co2 gas is called carbon fixation.
True
Lipids, proteins, and sugars all may serve as substrates of glycolysis.
False
Which of the following is true of catabolism?
Catabolism reactions are exergonic; they break down complex organic molecules into simpler ones.
Which of the following statements is true about enzymes?
Each enzyme has a characteristic three- dimensional shape.
The chemical reactions involved in synthesizing proteins and cell wall peptidoglycan are examples of ____ reactions.
anabolic
Which of the following statements is accurate concerning glucose metabolism?
Pyruvic acid, the product of glycolysis, is the starting block for both Krebs cycle and fermentation.
The complete oxidation (catabolism) of glucose typically involves three stages. The greatest amount of ATP is produced in which stage?
The elctron transport chain
Which of the following molecules carry electrons during various stages of glucose catabolism?
NADH & FADH2
Which of the following four stages of glucose oxidation requires molecular oxygen?
Oxidative phosphorylation
Which o fthe following statements accurately describes the difference between aerobic & anaerobic respiration?
Aerobic respiration use oxygen as the final electron acceptor & anaerobic respiration uses either an inorganic molecule, such as nitrate ions or sulfate ions, or an organic molecule, such as an acid or alcohol.
Carbon fixation occurs during______.
Photosynthesis
Fermentation differs from anaerobic respiration in all of the following ways EXCEPT that fermentation does NOT ___.
use an electron transport chain.
When fermentation test are used to help identify bacteria, which of the following end-products is typically detected bya color or change?
Acid
Which of the following are products of light -dependaent ( in italics) photosynthetic reactions & are necessary for the light- independent (in italics) photosynthetic reactions occur?
ATP& NADH
During the light-independent reactions of photosynthesis, organisms ____.
Synthesize sugars
Nitrobacter (italics) bacteria use carbon dioxide for their source and Nitrate ions as an energy source. This organism is a ____.
Chemoautotroph
Which group of organisms has members representing each of the following nutrional classification: chemoheterotrophs, chemoautogrophs, photoheterotrophs,&photoautotrophs?
Bacteria
Which of the following molecules traps energy released during oxidation reduction reactions?
ATP
Some amino acids are synthesized by adding an amine group to pyruvic acid or to one of the Krebs cycle intermediates. This process is known as ____.
Amination
You inocculate a bacterial culture into a tube containing glucose & peptides. The pH indicator shows that the pH decreased after 24 hours & then increased at 48 hours. Ehat has caused the increase in pH?
Deamination
Which of the following terms refers to pathways that can function both in anabolism & catabolism?
Amphilbolic pathways
Like glucose, amono acids are catabolized for energy, but these must be converted to a form where they can enter the Krebs cycle for oxidation. All of the following reactions occur in the catabolism or amino acids EXCEPT____.
carbon fixation
Microbial Growth
Increase in number of cells, not cell size
The Requirements for Growth
*Physical Requirements
- Temperature
-pH
-Osmotic pressure
*Chemical Requirements
-Carbon
- Nitrogen, Sulfure, and Phosphorous
- Trace elements
-Oxygen
-Organic Growth factor
Physical Requirements
* Temperature
-Minimum
-Optimum
-Maximum
Figure 6.1 Typical Growth rates of different types of microorganisms
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Psychrotrophs
Grow between 0 degrees Celcius and 20-30 degrees C
-cause food spoilage
pH
Most bacteria pH 6.5 and 7.5
Molds and yeasts pH 5 and 6
Acidophils grows in acidic environments
Osmotic Pressure
Hypertonic environments, increase in salt or surgar, plasmolysis
Extreme or obligate halophiles require osmotic pressure ( or they die)
Facultative halophiles tolerate
Chemical Requirements
* Carbon
-structural organic (has to have carbon in it) molecules, energy source
-Chemoheterotrophs use organic carbon sources
- Autotrophs use CO2
* Nitrogen
-In amino acids and proteins
-decompose proteins
-use NH4+or No3-
- few use N2 in nitrogen fixation
*Sulfur
* Phosphorus
In amino acids, Thiamine, and biotin
-decompose proteins
- use SO42- or H2S
* DNA, RNA, ATP, and membranes
* PO43-
Trace elements
Inorganic elements
* enzymes cofactors
Organic Growth Factors
-Organic Compounds obtained from the environment
- Vitamins, amino acids, purines and pyrimidines
Biofilms
Microbial communiites
-Form slime or hydrogels
- attracted by chemicals quorum sensing
Advantages of Biofilms
share nutrients
sheltered
Biofilms
indwelling catheters
low to cause infection
after exposure, patients developed infections
Agar
*Complex polysaccharide
-soildifying agent
-plates, slants, and deeps
* not metabolized by microbes
-liquefies at 100 degrees C
- Solidifies at -40 degree C
Culture Media
Culture medium: growth
Sterile: no living microbes
Inoculum: introduction of microbes
Culture: microbes growing
Chemically defined media: exact composition is known
Complex media: extracts and digests of yeasts, meat, or plants
- nutrient broth
- nutrient agar
Anaerobic Culture Methods
* Reducing Media
Contain chemicals (thioglycolate or oxyrase) that combine O2.
Heated to drive off O2.
Capnophiles
Require high CO2 conditions
CO2 packet
Candle jar
Biosafety levels
BSL-1: No special precautions
BSL-2: lab coat,gloves,eye protection
BSL-3: biosafety cabinets to prevent airborne transmission
BSL-4: sealed, negative pressure
*Exhaust air is filtered twice
Differential Media
Distinguish colonies of different microbes
Selective Media
Suppress unwanted microbes, encourages desired microbes
Enrichment Culture
Encourages growth of desired microbes
Obtaining Pure Cultures
Contains only one species or strain
COLONY a population of cells arising from a single cell or spore or from a group of attached cells
Colomy = Colony-forming unit
Streak plate method: isolate pure cultures
Preserving Bacterial Culutres
Deep- Freezing -50C to -95C
LYophilization (freeze-drying): vacuum
Reproduction in Prokaryotes
Binary fission
Budding
Conidiospores (actinomycetes)
Fragmentation of filaments
Understanding the Bacterial Growth Curve
1.Lag Phase: Intense activity preparing for population growth but no increasein population
2. Log phase:logarithmic or exponential increase in population
3. Stationary phase: Period of equilibrium; microbrial deaths balance production or new cells.
4. Death phase: Population is decreasing at a logarithmic rate
measuring Microbial Growth
Direct Methods
* Plate counts
* Filtration
* MPN
* Direct microscopic count
Indirect Methods
* Turbidity
* Metabolic activity
* Dry weight
Terminology of Microbial Control
Sepsis: contamination
Asepsis: absence of significant contamination
* Aseptic surgery- prevent microbial contamination of wounds.
Cont. Terminology of Microbial Control
Sterilization: removing all microbial life
Commerical sterilization: killing C. botulinum endospores
Disinfection: removing pathogens
Antisepsis: removing pathogens from living tissue
Cont. Terminology of Microbial Control
Degerming: removing micobes from a limited area
Sanitization: lowering microbial counts (eating utensils)
Biocide/germicide: killing microbes
Bacteriostasis: inhibiting not killing microbes
Effectiveness of treatment
Depends on:
-number of microbes
-environment
-Time of exposure
-Microbial characteristics
Actions of Microbial Control Agents
Alteration of membrane permeability
Damamge to proteins
Damamge to nucleic acids
Heat
Thermal death point (TDP): lowest temperature, all cells in a culture are killed in 10 min
Thermal death time (TDT): time during all cells in a culture are killed
Decimal Reduction Time (DRT)
Minutes to kill 90% of a population
Moist Heat Sterilization
denatures proteins
Autoclaves steam under pressure
Pasteuization
Reduces spoilage
Equivalent treatments
- 63 C for 30 min
- High-temperature short-time: 72 C for 15 min
- Ultra-high-temperature: 140 C for <1 sec
- Thermoduric organisms survive
Dry Heat Sterilization
* Kills by oxidation
- Dry heat
- flaming
- incineration
- Hot-air sterilization
Filtration
Hepa remmoves microbes
Mebrane filtration removes microbes
Physical Methods of Microbial Control
Low temperature : Inhibits growth
-Refrigeration
- Deep-Freezing
- Lyophilization
High Pressure: denautres proteins
Desiccation: prevent metabolism
Osmotic pressure: causes plasmolysis
Radiation
Ionizing radiation: x-rays, gamma rays, electrom beams
Nonionizing radiation: UV, 260nm
Microwaves: kill by heat, not especially antimicrobial
Principle of Effective Disinfection
Concentration of disinfectant
Organic matter
pH
time
Use-Dilution Test
Metal rigs dipped in test bacteria are dried
placed in disinfectant for 10 min at 20 C
transferred to culture media to determine bacteria survued treatment