Micro biology ch 8,7, 6

Metabolism
Sum of all chemical reactions within a living organism
Anabolism
building of complex organic compounds from simplier compounds
Catabolism
breakdown of complex organic compounds into simpler compounds
Anabolism
– Biosynthesis
-Covalent bonds are formed
-Dehydration synthesis
-Endergonic: more energy consumed than produced (requires ATP)
Ex. Nucleotides are used to synthesize nucleic acids.
Amino acids are used in the synthesis of proteins.
Endergonic
more energy consumed than produced (requires ATP)
Catabolism
-Degradative
– Covalent bonds are broken
-Hydrolytic reaction
Exergonic: more energy produced than consumed (produces ATP)
-Drive anabolic reactions
Ex. Complex sugars ? monosaccharides
Sugar ? CO2 and H2O
ATP
-High energy molecule
-Last 2 phosphate bonds are unstable
-Energy is released when last phosphate bond is broken
Metabolic Pathways
-Usually, a single covalent bond is made/ broken in each reaction.
-Begins with a specific molecule and ends with a final product (endproduct)
-Many steps are needed
-Process is efficient!
Collision Theory
All atoms, ions, and molecules are continuously moving. When they collide, electrons aredisrupted and can form/break chemical bonds.
Collision Theory
What factors determine if a reaction happens?
Velocity
Activation energy
Chemical configuration
Reaction Rate
-Frequency of collisions containing enough energy to result in a reaction
-Dependent on # of reactant molecules at/above activation energy
Temperature
Pressure
Enzymes
-Proteins that catalyze/speed up biochemical reactions without being permanently altered
Some are RNA
-Characteristic 3D shape determines function
substrate
-substance acted on by an enzyme
-Specificity: act on only one substrate
Active site:
region of enzyme that interacts with substrate
“Induced fit” between the substrate and the active site
-Not a lock and key fit
Enzyme Speed and Location
Number of substrate molecules converted per individual enzyme per second
1,000 – several million molecules per second!
How do enzymes work?
-Catalyze reactions by increasing probability of reactions
-Lower activation energy
-Increase # of molecules that reach activation energy
Ions: Fe2+,. Zn2+, Mg2+, Ca2+
Form bridge between substrate and enzyme
Coenzyme:
organic cofactor
Remove a chemical group from one substrate and transfer/carry it to another
H+, CO2, amino groups, or electrons
Electron carriers (accept and donate electrons)
Niacin
NAD+ (catabolic rxns), NADP+ (anabolic rxns)
Other B vitamins
FAD, Coenzyme A (important in Krebs cycle)
Electron carriers
accept and donate electrons
Factors Affecting Enzyme Activity
Temperature
pH
Substrate Concentration
Presence of Inhibitors
Temperature
Increasing
Denaturation
Loss of protein structure
Optimum temp
– max enzyme activity
– 35-40oC for most pathogens
pH Denaturation
-Loss of protein structure

-H+ and OH- compete with ions
in ionic bonds

Optimum PH
pH at which enzyme activity is maximum
Substrate Concentration
Increases in substrate concentration increase
enzymatic activity until all of the active sites are full
Saturation
-All active sites are full
-Curve levels off
Feedback inhibtion
Common way to regulate metabolic pathways
Oxidation
loss of electrons
Reduction
gain of electrons
Redox reaxtions
Sometimes a electron is exchanged with a proton(H+)
Metabolic Pathways
-Series of connected chemical reactions within cell
-Enzymes at every step
-Store & release energy from organic compounds
Carbohydrate catabolism
-Common strategy for microbes to make ATP
-Carbs are easily oxidized (good fuels)
Glucose C6 H12 06
-Highly reduced molecule
-Lots of energy
Cellular Respiration
ATP-generating process in which molecules are oxidized and the final electron acceptor is an inorganic molecule
Aerobic
oxygen
Anaerobic
Anaerobic = not oxygen
Glycolysis
-sugar splitting
-1st stage of carb catabolism
Glycolysis
Used in most living cells
-Some bacteria have alt. pathways
Glycolysis
-Almost every step requires an
enzyme
Glycolysis
Splits 6C sugar into 2 3C molecules
Glycolysis
Oxidation & rearrangement to form
pyruvic acid
Glucose
2 pyruvic acid
(pyruvate is ionic form)
Glycolysis
Net gain = 2ATPs + 2NADH
Glycolysis
-Does not require oxygen
-Followed by respiration or
fermentation
krebs cycle
Gradually releases energy through e- transfer
krebs cycle
-E- transferred to carrier coenzymes

-NAD+ and FAD
(reduced to NADH and FADH2)

krebs cycle
Net 2 ATP + 6NADH + 2FADH2
*Per glucose molecule*
krebs cycle
Citric Acid Cycle
Tricarboxylic Acid Cycle
Electron Transport Chain
Several types of ETCs
– Even in same organism
-All function similarly
Electron Transport Chain
Location
-PM of bacteria
-Inner mitochondrial membrane of euk.
Electron Transport Chain
Carriers transfer e- energy into ATP
-Series of redox reactions
proton motive force
Energy drives H+ pumps creating
chemiosmosis
In the ETC H+ flow through ATP synthase
ATP Synthase
NADH3—>ATP
FADH2—>2ATP
Prokaryotes
38 ATPs from one glucose molecule
Eukaryotes
-36 ATPs
-Energy lost with transfer of NADH/FADH2 across membrane
Anaerobic Respiration Summary
-final e- acceptor = inorganic molecule (not O2)

– 2-36 ATPs / glucose

Anaerobic Respiration
2-36 ATPs / glucose
Eukaryotes
36 ATPs
Fermentation
Releases energy from sugar or other organic molecules
-Incomplete oxidation of carbohydrate
-Final e- acceptor is organic molecule
Fermentation
Final e- acceptor is organic molecule
Fermentation
-Oxygen not required (anaerobic)
-CAN occur in presence of O2
-2 ATPs made during glycolysis
– Krebs cycle and Electron transport not used
Fermentation
Glycolysis followed by pyruvate reduction
-NADH passes e-s to pyruvate instead of ETC
Fermentation
Endproducts depend on microbes, substrates, enzymes
Alcohol Fermintation
Ethanol produced
Acid Fermintation
Lactate
Carbon
50% dry weight of a bacterial cell
Carbon
obtain through energy sources (carbs,lipids,proteins)or Co2
Carbon
4 nutrional caegories
-Photoautotroph
-Chemoautotroph
-Photoheterotroph
-Chemoheterotroph
Hydrogen
-In all organic and most inorganic compounds
-Maintains pH, forms hydrogen bonds, source of free energy in redox reactions
Hydrogen
-Maintains pH, forms hydrogen bonds, source of free energy in redox reactions
Hydrogen
-In all organic and most inorganic compounds
Nitrogen
Needed for protein, nuceic acid, and ATP synthesis
Nitrogen fixation
conversion of N2(in air) into NH4
Phosphorous (P)
-DNA& RNA
-Phospholipids
-ATP
Sulfur(S)
Needed for protein (in Cys and Met)& vitamin synthesis (Biotin & Thiamine)
Apoenzyme
-protein portion
-inactive
coenzyme
cofactor
-non protein portion
-activator
Holoenzyme
-Whole enzyme
-active
Ions
Potassium, sodium, magnesium,calcium, sodium
Essential organic molecules
growth factors
Aerobes require
– O2 for aerobic respiration
– H+ (stripped from organic compounds) + O2(air) + electrons—->H2O
Anaerobes
-don’t use 02 for energy production
-Anarobic respiration & fermentation
radicals
toxic products that oxygen is transferred into while it is being utilized
Radicals are formed when
– when o2 is used
-by ionizing radiation (H2O–>H+ (+) OH-)
Oxygen radicals
O2-:superoxide
H2O2:Hydrogen peroxide
OH-:hydroxyl ion
O2^2:Peroxide ion
O2-
superoxide
H2O2
Hydrogen peroxide
OH-
hydroxyl ion
O2^2
Peroxide ion
Why are oxygen radicals toxic?
– They are unstable
-Disrupts eletron arrangements around other molecules
-causes DNA or cell damage
Detoxifaction
-naturally occurring antioxidants
-Detoxifying enzymes
Naturally occurring antioxidants
-Tocopherol(vitamin E)
-Ascorbic acid (Vitamin C)
-Melatonin
Detoxifying enzymes
-Superoxide dismutase
-Catalase
-Peroxidase
Obligate Aeorobes
only aerobic growth;oxygen required
-Medium:Growth occurs only where high concentrations of oxygen have diffused into the medium
-Presence of enzymes catalase and superoxide dismutase (SOD) allows toxic forms of oxygen to be neutralized;can use oxygen
Facultative Anaerobes
-Both aerobic and anaerobic growth; greater growth in presence of oxygen
-Medium:Growth is best where most oxygen is present, but occurs throughout tube
-Lacks enzymes to neutralize harmful forms of oxygen; cannot tolerate oxygen
-Presence of enzymes catalase and SOD allows toxic forms of oxygen to be neutralized;can use oxygen
obligate anaerobes
-organisms that survive in habitats without oxygen
-only anaerobic growth; ceases in presence of oxygen
Medium:Growth occurs only where there is no oxygen
Aerotolerant Anaerobes
-Medium:growth occurs evenly; oxygen has no effect
Presence of one enzyme, SOD, allows harmful forms of oxygen to be partially neutralized; tolerates oxygen
Microaerophiles
-Only aerobic growth; oxygen required in low concentration
-Medium:Growth occurs only where a low concentration of oxygen has diffused into medium
-Produce lethal amounts of toxic forms of oxygen if exposed to normal atmospheric oxygen
In aerobic respiration carbohydrates are ultimately broken down into
CO2
chemiosmosis
Most ATP produced in aerobic respiration occurs in the process of
pyruvate
In glycolysis the most reduced compound formed is
ATP
In glycolysis, the activation of glucose is accomplished by:
oxygen
The final electron acceptor in aerobic respiration is
glycolysis
Which stage of aerobic respiration requires ATP
none
. Which stage of aerobic respiration requires CO2?
ATP synthase
As protons flow through the ______, energy is released and exploited to combine ADP and inorganic phosphate to form ATP.
ATP synthase
is an enzyme, a molecular motor, an ion pump, and another molecular motor all wrapped together in one amazing nanoscale machine. It plays an indispensable role in our cells, building most of the ATP that powers our cellular processes
-Prokaryotes
38 ATPs from one glucose molecule
-Eukaryotes get 36 ATPs
Energy lost with transfer of NADH/FADH2 across membrane
Summary of Aerobic Respiration
Anaerobic Respiration
-final e- acceptor = inorganic molecule (not O2)
– 2-36 ATPs / glucose
Fermentation
-Releases energy from sugar or other organic molecules
-Incomplete oxidation of carbohydrate
-Final e- acceptor is organic molecule
Fermentation
-Oxygen not required (anaerobic)
-CAN occur in presence of O2
– 2 ATPs made during glycolysis
– Krebs cycle and Electron transport not used
Fermentation
-Glycolysis followed by pyruvate reduction
-NADH passes e-s to pyruvate instead of ETC
Fermentation
Endproducts depend on microbes, substrates, enzymes
True
Fermentation can occur in the presence of oxygen
50% of dry weight of a typical bacterial cell
Carbon
Carbon
Obtain through energy sources (carbs, lipids, proteins) or CO2
Carbon
4 Nutritional categories
Photoautotroph
Chemoautotroph
Photoheterotroph
Chemoheterotroph
Photoautotroph
are organisms that carry out photosynthesis. Using energy from sunlight, carbon dioxide and water are converted into organic materials to be used in cellular functions such as biosynthesis and respiration.
Chemoautotroph
Chemoautotrophs are organisms that obtain their energy from a chemical reaction (chemotrophs) but their source of carbon is the most oxidized form of carbon, carbon dioxide (CO2).
Photoheterotroph
Photoheterotrophs depend on light for their source of energy and mostly organic compounds from the environment for their source of carbon.
Chemoheterotroph
are unable to fix carbon to form their own organic compounds
Hydrogen
In all organic and most inorganic compounds
Maintains pH, forms hydrogen bonds, source of free energy in redox reactions
Nitrogen
Needed for protein, nucleic acid, and ATP synthesis
Nitrogen fixation
conversion of N2 (in air) into NH4+
Phosphorous (P)
is an essential element in the formation of phospholipids, a class of lipids that are a major component of all cell membranes, as they can form lipid bilayers, which keep ions, proteins, and other molecules where they are needed for cell function, and prevent them from diffusing into areas where they should not be. Phosphate groups are also an essential component of the backbone of nucleic acids and are required to form ATP – the main molecule used as energy powering the cell in all living creatures.
Sulfur
Needed for protein (in Cys & Met) & vitamin synthesis (Biotin & thiamine
Ions:
Potassium, sodium, magnesium, calcium, sodium
Aerobes
-require O2 for aerobic respiration

-H+ (stripped from organic cmpds) + O2 (air) + electrons? H2O

Anaerobes
don’t use O2 for energy production
radicals
As oxygen is utilized it is transformed into several toxic products
Radicals are formed
-when O2 is used
-by ionizing radiation (H2O ? H+ + OH-)
-They are unstable
– Disrupt electron arrangements around other molecules
Why are oxygen radicals toxic
obligate anaeorobe
Which of the following cannot survive in the presence of O2?
Solute
– substance dispersed within a solvent
-solid sometimes gas
Solvent
– dissolving medium (usually liquid)
-usually water, or a liquid
Solution
– mixture of substances that cannot be separated by filtration
-combinationation of substances that solvents are dissolved
Concentration gradient
Difference in concentration on two sides of a membrane
Passive
moves from high to low conc;
no energy/ATP required
Active
moves from low to high conc;
energy required
Passive Processes
Simple Diffusion

Facilitated Diffusion

Osmosis

Simple Diffusion
Net movement of solute molecules —from high to low concentration
-Stops at equilibrium
Facilitated Diffusion
s the process of spontaneous passive transport of molecules or ions across a cell’s membrane via specific transmembrane integral proteins.
Osmosis
Net movement of solvent (usually H2O) across selectively permeable membrane
Passive transport of solvent
Movement follows the CG for solvent
simple diffusion
once the molecules stop at equilibrium they dont stop moving
small uncharged molecules
what kind of molecules can easily go through simple diffusion
Facilitated Diffusion
charged molecules go from high to low. They cant cross the membrane on their own because they are charged so they need carrier proteins transport proteins and permeass to act as channels
Osmossis
-most instance water is the solvent
-water doesnt move easiliy or quickly because it has a partial charge
Aquaporin
allows more rapid, faster movement of water
Isotonic
-is equal, still has movement inside and outside
-rates of diffusion are equal in both directions
Hypotonic
less solute outside the cell compared to the inside
cell wall:the cell swells, the cell wall resists osmotic pressure prevents it from bursting
no cell wall: cell swells and may burst if no mechanism exists to remove water
Hypertonic
Higher concentration outside
Cell wall: water diffuses out of the cell and shrinks the cell membrane away from the cell wall
no cell wall: water diffuses out of the cell causes it to shrink and become distorted
Gram +
Which bacteria are more susceptible to osmotic lysis?
Active Transport
-Require ATP
-Usually move against CG
Can be with CG but at a faster rate

-Requires carrier protin
-Examples
*Ion pumps
*Group Translocation
Transport is coupled with alteration
Only in prokaryotes
*Endocytosis

Group Translocation
-Transport is coupled with alteration
-Only in prokaryotes
Endocytosis
is a form of active transport in which a cell transports molecules (such as proteins) into the cell (endo- + cytosis) by engulfing them in an energy-using process.
Phagocytosis
process by which certain living cells called phagocytes ingest or engulf other cells or particles. The phagocyte may be a free-living one-celled organism, such as an amoeba, or one of the body cells, such as a white blood cell.
Pinocytosis
the ingestion of liquid into a cell by the budding of small vesicles from the cell membrane.
Receptor-mediated
is a process by which cells absorb metabolites, hormones, other proteins –
Acidophiles
acid-tolerant microbes
Stomach pH = 1 (Helicobacter pylori)
Vagina pH = 3-5 (Lactobacillus)

Acids from fermentation preserve foods

Microbial wastes are acidic
Use buffers in media

Hypotonic Solutions
If concentrations of dissolved solutes are less outside the cell than inside, the concentration of water outside is correspondingly greater. When a cell is exposed to such hypotonic conditions, there is net water movement into the cell. Cells without walls will swell and may burst (lyse) if excess water is not removed from the cell. Cells with walls often benefit from the turgor pressure that develops in hypotonic environments.
Reducing media
contains ingredients that deplete O2
Ex. Thioglycollate medium (liquid)
Anaerobic jars
Used for plate cultures
Colony growth
Capnophiles
Require CO2 & some O2
-Intestines, respiratory tract, body tissues
– CO2 incubator or candle jar
Biofilms
Benefits
-Share nutrients
-Shelter
?-opportunity for genetic exchange
Cooperative activity
Quorum sensing
Communication via chemical cues
Bacterial Growth
Most reproduce by binary fission
Budding
Outgrowth of cell that grows and
separates from parent
External Spores (conidiospores)
– Filamentous bacteria
– Spores develop into new cells
-More common for fungi
Fragmentation
New cells grow from fragments
Bacterial Growth
Most culturable bacteria reproduce by binary fission
Generation Time
Time required for cell number to double
Lag Phase
-cell number not increasing
-metabolic activity is still going on
Log Phase
-Increase in population of bacteria cells
-Exponential growth
-shortest generation time
-Greatest Growth
Stationary Phase
-Nutrient depletion
-slows growth
-cell division=cell death
-waste product
-period of equilibrium
Death Phase
-still cell division is occurring
-death is occurring faster rate than growth
-now death out weighs growth
lag phase
In lab, you are asked to inoculate a Phenol Red broth with E. coli. You immediately place the tube in the 37?C incubator. In which phase of the bacterial growth curve is your culture?
death phase
You leave your culture in the 37?C incubator for five days. In which phase is the culture most likely in when you remove it?
Direct measurement
-Technique that involves some counting
– Count small sample & determine total population
Indirect measurement
-Estimation of cell numbers without counting cells
– Turbidity, Mass, Spectrophotometry
Plate Counts
Direct count

-Estimates live cells by counting colonies
-30-300 CFU/plate
-Pour plate method or spread plate method

Filtration Method
-Volume passed through membrane (bacteria trapped)
– Transfer to plate and grow
-Count colonies and divide
by number of ml filtered (CFU/ml)
-Good when bacteria # is small
Can’t count 1CFU/ml
Ex. Water samples
(coliform bacteria)
Direct Microscopic Count
Count number of cells spread over grid using a microscope

-Advantages: quick (no incubation required)
counts all cells (live and dead)
-Disadvantage: difficult to count motile bacteria

Turbidity
-Indirect
-Uses spectrophotometer
Measures amount of light
passing through a sample
% transmittance
Absorbance
Turbidity
-Advantage: quick and easy

-Disadvantages:
must be read during log phase
needs to be correlated to direct measurement

Turbidity Method
-Indirect method of measurement
-Degree of cloudiness, turbidity, reflects the relative population size
-Advantage: quick and easy
-Disadvantages:
must be read during log phase
needs to be correlated to direct measurement
Direct microscopic count
You are part of the Quality Control team that measures the microbial growth of Horizon milk. You are expected to deliver same-day results before the milk is shipped. Which of the following techniques do you most likely use for your job?
Hypertonic Solutions
If concentrations of dissolved solutes are greater outside the cell, the concentration of water outside is correspondingly lower. As a result, water inside the cell will flow outwards to attain equilibrium, causing the cell to shrink. As cells lose water, they lose the ability to function or divide. Hypertonic environments such as concentrated brines or syrups have been used since antiquity for food preservation because microbial cells that would otherwise cause spoilage are dehydrated in these very hypertonic environments and are unable to function.
General Characteristics of Viruses
-Not all have spikes
-Not all cause disease
-Requires electron Microspy
-All Acellular
-Either DNA or RNA genome
-Have to have a host cell
-Small amounts of DNA or RNA
-All have a protein capsid
Virion
-fully developed virus particle, infectious
-if feature is missing it is not a virion
Virus Genome
SS DNA

DS DNA-our genome

SS RNA

DS RNA-never seen in our genome unless infected by a virus

capsid
protect nucleic acids
Envelope
-not all viruses have a envelope
– around capsid
-came from host cell usually the PM from the host
-it has a envelope because it is most likely to dry out
Influenza
-8 segmented genome
-Hemaglutin is the attachment
-Affect Respiratory system
– an RNA virus
RNA Viruses
Go through Antigentic drift
capsomeres
Virus Protein coat is made of
nucleocapsid
-Protects nucleic acid
-Capsid + nucleic acid =
Icosahedral or helical shape
Virus structure
Viral structure:Enzymes
All viruses use some host machinery
Ribosomes
DNA polymerases
Drugs that interfere with these enzymes disrupt viral AND host cell function
All RNA viruses need an RNA polymerase
We can target these with drugs!
Some Viruses package their own (viral) enzymes which is
Enveloped viruses
-Phospholipid membrane around capsid
From host plasma or nuclear membrane

-Must stay moist during spread from host to host

-Viral proteins and sugars bind host receptors
-Attachment factors on virion surface
Naked viruses
Enveloped viruses
-Determine host range of virus
Ex. Influenza & HIV
Viral Attachment Factors
-viruses dont have a cell structure
-No domains
Why aren’t viruses classified according to the same taxonomic system as prokaryotic and eukaryotic microbes?
Determines how it gets copied
Virus Strategy for replication
Obligate intracellular parasites

Host-specific

Can be cultured only inside living cells
Bacteria
Living animals
Chicken egg
Cultured cells

Growth of Viruses
bacteriophage
Which of the following is a virus that infects bacteria?
Virion
infective form
Lytic cycle
multiplication in host cell in which host cell is disrupted to release virions
Lysogenic cycle or latent phase
Dormant stage when virus is not undergoing “active” replication; host cell is not killed
Lytic cycle
Which type of viral life cycle causes the host cell to die?
lysozyme
Which of the following enzymes is necessary for T4 phage entry into and exit from a host bacterial cell like E. coli?
Attachment
Bacteriophage:Tail fibers attach to cekk wall proteins

Animal virus:Attachment sites are plasma proteins and glycoproteins

Penetration
Bacteriophage:viral DNA injected into host cell

Animal virus:CApsid enters by endocytosis or fusion

Uncoating
Bacteriophage:not required

Animal virus:Enzymatic removal of capsid proteins

Biosynthesis
Bacteriophage:in cytoplasm

Animal virus:In nucleus (DNA viruses) or cytoplasm (RNA viruses)

Chronic Infection
Bacteriophage:Lysogeny

Animal virus:Latency; slow viral infections; cancer

Release
Bacteriophage:Host cell lysed

Animal virus:Enveloped viruses bud out; nonenveloped viruses rupture plasma membrane

Influenza Virus
-Enveloped RNA Virus
– Hemaglutinin and neuraminidase determine Influenza strains
Antigenic Shift
Pigs susceptible to avian and human flu strains
Segmented genome allows mixing in pigs (intermediate host)
-RNA remains in the cytoplasm
-All RNA viruses make an RNA-dependent RNA polymerase
Copies RNA strand
(+ sense / – sense strands)
-Translation by host
-Virus Assembly
Animal Viral Biosynthesis: RNA Genomes
-Viral DNA moves to the nucleus
-DNA is transcribed
-Translation by host ribosomes
-DNA is replicated in nucleus
-Virus assembles
Animal Viral Biosynthesis: DNA Genomes

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