MICROBIAL METABOLISM ● Occurs through a series of

enzyme-catalyzed reactions ​that forms

● Production and utilization of energy
Living Organisms​ - divided according to the
chem form in which they obtain C from
environment
● Autotrophs - photosynthetic bacteria and
vascular plants
○ Uses CO2 from atmosphere as
sole source of C
○ Most are photosynthetic, obtain
energy from sunlight
● Heterotrophs
○ Can’t use atmospheric CO2
○ Obtain C from their environment in
form of complex organic molecules
(ex. glucose)
○ Survive on the products of other
organisms
● C, O2, and H2O are constantly cycled
between heterotrophs and autotrophs
Cellular Respiration
Organisms and Energy
metabolic pathways
○ Catabolism
■ Oxidation ​(broken down)
■ Degradative phase of
metabolism
■ Organic nutrient molecules
are converted to ​smaller
and ​simpler ​end products
■ Releases ​energy
○ Anabolism
■ Reduction or Biosynthesis
(built up)
■ Small simple precursors are
built ​up into l​ arger more
complex m ​ olecules
● Lipis
● Polysaccharides
● Proteins
● Nucleic acids
● Production of ​ATP ​whose molecules are
oxidized​ to produce energy
● ADP ⇝ ATP → ADP → release of energy
○ Aerobic Respiration
■ Occurs in bacteria living in
oxygenated e ​ nvironments
■ Oxygen ​- final electron
acceptor in reaction
○ Anaerobic Respiration
■ Occurs in bacteria living in
oxygenated environments
■ Oxygen kills bacteria
■ Inorganic molecule​ - final
electron acceptor in
reaction

Glucose
● The most important carbohydrate
● Autotrophs and Heterotrophs are
● Considered the ​energy-currency m​ olecule
dependent on each other
for autotrophs and heterotrophs
● Three major outcomes:
Metabolism
● Is the sum of all chemical transformation
that occurs in a cell or organism
 Substrate Product Enzyme and
(Cofactor)

1. Glucose * G-6-P Hexokinase

(Mg²⁺)

2. G-6-P F-6-P Phosphohexose

Isomerase

3. F-6-P * F-1,6-BP PFK-1 (Mg²⁺)

4. F-1,6-BP G-3-P Aldolase

5. G-3-P DAP TPI

○ May be ​stored ​(polysaccharide or
sucrose) 6. G-3-P 1,3-BPG G-3-PD (Mg²⁺)
■ Glycogen - a ​ nimals
■ Starch -​ plants 7. 1,3-BPG 3-PG PGK (Mg²⁺)
○ May be ​oxidized ​to a ​three-carbon
8. 3-PG 2-PG PGM
compound (pyruvate) via ​glycolysis
■ Provide ​ATP​ and​ metabolic 9. 2-PG PEP Enolase (Mg²⁺)
intermediates
○ May be ​oxidized ​via ​pentose 10. PEP * Pyruvate PK (Mg²⁺)
phosphate pathway y​ ielding:
■ Ribose r-phosphate ​for * = Irreversible
nucleic acid synthesis
■ NADPH ​for reductive ● Acetyl-CoA Formation and Citric Acid
biosynthetic processes Cycle or Kreb’s Cycle (​Mitochondria​)

Substrate Product Enzyme

1. Acetyl CoA Citrate Citrate

& oxaloacetate Synthase

2. Citrate Isocitrate Aconitase

3. Isocitrate * AKG Isocitrate

Dehydrogenase

4. AKG Succinyl CoA AKGD complex

5. Succinyl Succinate Succinate

CoA Thiokinase

6. Succinate Fumarate Succinate

Dehydrogenase

7. Fumarate Malate Fumarase

8. Malate Oxaloacetate Malate

Three Stages of Aerobic Metabolism Dehydrogenase
● Glycolysis (​Cytoplasm)​
* = Irreversible
 ● Electron Transport or Oxidative
Glycolysis Cytoplasm Cytoplasm
Phosphorylation (​Mitochondria​)
Intermediate Cytoplasm Cytoplasm
Step
Complex
Kreb’s Cycle Mitochondrial Cytoplasm
Complex I: NADH ● Rotenone
Matrix
Dehydrogenase amytal - blocks
electron ETC Mitochondrial Plasma
transfer from Inner Membrane
Complex 1 to Membrane
CoQ
● Removes H in Overview of Anaerobic Metabolism
NADH
● Energy in ​glucose​ is released ​without the
Complex II: Succinate ● Removes H presence of oxygen
Dehydrogenase from succinate ● Lactic acid ​- end product of anaerobic
● Observed in metabolism
kreb’s cycle ○ Recycled back by the ​liver​ in
humans
Complex III: ● Coenzyme Q
● Pyruvate → Lactate
Ubiquinone - transfers
○ Enzyme: Lactate Dehydrogenase
Cytochrome C electrons from
Oxidoreductive Complex II to
ATP Production Requirements
III
● Antimycin A -
blocks electron
transfer with
complex III

Complex IV: ● Chemiosmotic

Cytochrome Oxidase Theory - H+
gradient drives
ATP Synthesis

● Glucose + O2 = CO2 and H2O

Respiration and Fermentation

● Respiration
○ Glycolysis
○ Acetyl-CoA Formation
○ ETC
● Fermentation Questions
○ Pyruvate → Acetaldehyde →
Ethanol ● Which cellular respiration pathway is the
most efficient in producing ATP?
Carbohydrate Catabolism ○ A: ​Aerobic Respiration
● What kinds of human tissues do you think
Pathway Eukaryote Prokaryote anaerobic live in?
 ○ A: ​Parts of the human body not ● Obtained from water
exposed to oxygen (ex. bladder) ■Carbon
● What about aerobic bacteria? ● Backbone for
○ A:​ Parts of the body with oxygen organic materials
present (ex. respiratory tract - ● Make up 50% of dry
pneumonia ) weight of bacteria
● Where does the electron transport chain ● Heterotrophs -
occur in eukaryotic cells? organic materia
○ A: ​Inner Mitochondria ● Autotrophs - CO2
● What about in prokaryotic cells? ■ Nitrogen
○ A: ​Inner Plasma Membrane ● Earned by
decomposing
MICROBIAL GROWTH proteins or inorganic
nitrogen sources
Growth Requirements ■ Sulfur
● Physical ● Synthesize sulfur
○ Temperature containing amino
■ Mesophiles - moderate acids
■ Psychrophiles - cold ○ For proteins
■ Thermophiles - heat and vitamins
● From sulfur
containing amino
acids or ions
■ Phosphorus
● Synthesizes nucleic
acids and
phospholipids in cell
membrane and ATP
○ Trace elements
■ Copper
■ Molybdenum
○ pH ■ Zinc ions
■Around neutral ○ Organic growth factors
■Some bacteria produces ■ Organic compounds that
acidic waste products can’t be synthesized
○ Osmotic Pressure ● Vitamins
● Some amino acids
● Purines and
pyrimidines
Oxygen Requirements

■ Isotonic - ideal
■ Hypertonic - crenation
■ Hypotonic - lysis
● Chemical Requirements
○ Major elements
■ Hydrogen and Oxygen
 ● Obligate aerobes - require oxygens
● Facultative anaerobe - can grow
aerobically or anaerobically
● Obligate anaerobe - cannot grow in the
presence of oxygen; does not have
catalase (SOD - superoxide dismutase)
● Aerotolerant anaerobes - can survive with
oxygen but cannot used it
● Microaerophiles - only require or tolerate
small concentrations of oxygen
Culture Media
● Chemically defined - exact contents are
known; for fastidious organisms
● Complex - extracts from yeast
● Reducing - substances that removes
oxygen; for anaerobe
● Selective - suppresses the growth of
unwanted and encourage the growth of
desired microbes; for clinical studies
● Differential - distinguish between two types
of microbes; allows identification of
bacteria; ex. Blood agar
● Enrichment culture - for hard to grow
microbes; target specific microbes that are
finicky
Division of Bacteria: Binary Fission
● Asexually
● Steps:
○ Bacteria copy its chromosome
○ Elongates
○ Splits of into two daughter bacteria
■ Identical to the parent
bacteria
● Process can be continuous and is rapid
Growth Phase
● Lag Phase
○ Dormant bacteria is transferred to a
fresh medium with nutrients
○ Metabolic state takes time to switch
from dormant to active state
○ Less than an hour to days
● Log Phase
○ Bacteria is actively undergoing
binary fission
○ Bacteria double numbers every
generation period
■ Ranging fro​m 20 mins. - 2
days
○ Plenty of nutrients​ and​ little waste
build up​ will help the bacteria grow
continuously
● Stationary Phase
○ Nutrient depletion occurs and some
bacteria begin to die
○ Number of new cells = number of
cells that are dying
● Death Phase
○ Bacteria has depleted most of its
nutrients
○ Waste build up increases
○ Number of bacteria that die are
larger than they are made
○ If carried on most of the cells die
○ If cell wants to survive metabolism
should be switch to dormant state
CONTROL OF MICROBIAL GROWTH
Terminology
● Began in the 18th century
○ Louis Pasteur
○ Robert Koch
○ Joseph Lister
● Sterilization
○ Destruction or removal of forms of
microbial life, including endospores
○ Done before and after experiments
○ Usually done in the autoclave
■ ↑ pressure = ↑ temperature
■ 15 psi; 121 ℃
● Commercial Sterilization
○
● Disinfection
 ○ Destruction of vegetative ○ Bactericidal - not only prevents but
pathogens in ​inanimate objects kills the bacteria as well
■ Ex. lysol, bleach Antibiotic Modes of Action
● Antisepsis ● Targets specific metabolic processes
○ Destruction of vegetative ○ Replication
pathogens on ​living tissues ■ Quinolones
■ Ex. listerin ■ Not always used for first
● Degerming line defense since it has
○ Removal of microbes from a limited side effects
area ○ Transcription
■ Ex. skin around injection ○ Translation (Protein Synthesis)
● Sanitization ■ Chloramphenicol
○ Treatment intended to lower ■ Erythromycin
microbial counts on eating and ■ Tetracycline
drinking utensils to safe public ■ Streptomycin
health levels ■ Go after the ribosomes
■ Ex. hand sanitizer ○ Plasma Membrane Injury
■ Pokes hole in cell
Antibiotics membrane and creates
● Penicillin leaks
○ First antibiotic ○ Metabolism (Synthesis of essential
○ Alexander fleming metabolics)
○ Bacteria was inhibited by mold that ■ Metabolism that are not
was produced a certain compound found in eukaryotic cells
(Narrow Spectrum)
Spectrum of Antimicrobial Activity
Structure of Some Common Antibiotics
● Prokaryotic VS Eukaryotic
○ Cell Wall ● Ampicillin
■ Pro - ✓
■ Eu - x
○ Ribosomes - few difference that
makes antibiotic target prokaryotic
but not eukaryotic
○ Detail of Metabolisms
■ May target specific ● Erythromycin
pathways
● Broad Spectrum VS Narrow Spectrum
○ Depending on the range of
microbes they affect
○ Broad spectrum may affect all
gram-negative bacteria
○ Narrow may only work on a few
genera or species
● Bacteriostatic VS Bacteriocidal
○ Bacteriostatic - not necessarily kill
living cells but inhibits division and
growth
 ● Polymyxin B
● Tetracycline
Inhibition of Cell Wall Synthesis
● Peptidoglycan has alternating NAM and
NAG molecules held together by cross
bridges
● Penicillin prevents the formation of the of
the cross bridges
○ Makes the cell porous and cell
cannot resist osmotic pressure
making it rupture
○ Only affects cells that are trying
grow (Bacteriostatic)
Inhibition of Protein Synthesis
● Streptomycin
○ Changes shape of 30S component
of ribosome causing misreading of
mRNA code
○ Lead to production of protein that
was not encoded for
● Tetracyclines
○ Interfere with attachment of tRNA
to mRNA - ribosome complex
○ Get in the way of bringing the right
amino acids
● Chloramphenicol
○ Binds to 50s portion and inhibits
formation of peptide bond
Kirby-Bauer Test for Sensitivity to Antibiotics
● Also known as the “Disk Diffusion Method”
● Culture is spread on the plate and disk
impregnated with antibiotics at certain
concentrations are placed in the surface
● Shows if bacteria is sensitive or resistant
to the antibiotic through zone of inhibition
● Zone of Inhibition
○ Sensitive - bacteria growth is
inhibited or killed
○ Resistant - bacteria can grow in the
presence of the antibiotic
Broth Dilution and MIC Determination
● Helps in determining whether antibiotic is
either bacteriostatic or bactericidal
● May also be of help in prescribing
antibiotics to know if
○ Side effects
○ Causes secondary infection
● MIC
○ Minimum inhibitory concentration
○ Serial dilution of antibiotics +
culture
○ Measured in spectrophotometer;
absorbent in 600 nm
○ Where cloudiness (growth) drops
off
○ Not cloudy = no growth; minimum
concentration of antibiotic needed
○ Differs in each organism

■ Example
● S. aureus - lower
conc.
● E. coli - higher conc.
● MBC
○ Minimum Bactericidal
Concentration
○ Non-cloudy tubes may still contain
bacteria but they are not actively
dividing
○ Amount of concentration needed to
kill ​bacteria
○ Take culture from the tube, plate,
and incubate to observe if there are
still growth or cell still present
Antibiotic Resistance
● Four basic ways bacteria “combat”
effectiveness of antimicrobial agents:
○ Destruction or inactivation of the
drug
■ Ex. penicillin - bacteria
produces beta-lactamase
which breaks down
beta-lactam ring, a structure
of penicillin
○ Prevention of penetration to the
target site within microbe (cell wall)
■ Ex. Tetracycline - bacteria
allows it to enter but binds it
up inside which prevents it
from reaching the ribosome
○ Alteration of drug’s target sites
■ Ex. changes the shape of
the ribosome so that the
antibiotic cannot bind to it
○ Rapid efflux or pumping of the drug
out of the cell
■ Ex. when the antibiotic gets
in the cell it gets pump out
of the cell which stops it
from infecting the inside of
the cell
MICROBIOLOGY OF MICROBIAL GENETICS
Gene
● A portion of chromosome that determines
or affects a single character or phenotype
○ 1960s - thought to be a segment
of genetic material encoded for one
enzyme (one gene-one enzyme)
○ Recently broaden to one gene-one
polypeptide (sequence of amino
acid)
● Biochemical definition:
○ All the DNA that encodes the
primary sequence of final gene
product
■ Can either be polypeptide
​ r
or RNA with ​structural o
catalytic​ function
● Regulatory sequence
○ Provide signals that denote
beginning or end of genes
○ Influence transcription of genes
○ Act as initiation points for
replication or recombination
Genetic Code
● 3 base pairs - 1 amino acids
● Sequences determine the arrangement of
amino acids
● Amino acids are connected by polypeptide
chains
Transcription and Replication ● Translationof mrRNA begins at the start
codon
○ AUG
● Ends at nonsense codon
○ UAG
○ UGA
○ UAA

Gene Regulation
● Constitutive genes - expressed at fixed
rate
● Other genes - only expressed as needed
● Gene transfer can occur between ○ Repressible genes
individuals of the same generation ○ Inducible genes
● Way in which we can figure out if we
Replication of Bacterial DNA should continue synthesizing certain
proteins

● Active Repressor Protein - produced to

● Similar to eukaryotic replication
stop synthesizing protein
● Occurs in circular motion
● Inducer - lands on the repressor protein
and inactivate it
Transcription of Bacterial DNA
● Process where DNA ultimately produces
Germline Mutation
protein according to arrangement of
● Evolutionary Biology
nucleotide bases
○ Genetic changes to germ cells
○ DNA is transcribed to make RNA
■ Enhance or disrupt
■ mRNA
population’s fitness or
■ tRNA
ability to reproduce
■ rRNA
■ Only germ cell mutations
○ Transcription begins when RNA
are transmitted to offspring
polymerase binds to the promoter
● Molecular Biology
sequence
○ Permanent, transmissible change
○ Transcription proceeds in the 5’ ⟶
in the nucleotide sequence of
3’ direction
chromosome
○ Transcription stops when it reaches
○ Usually in a single gene
the terminator sequence
○ Leads to a change or loss of its
○ There is no nucleus
normal function
○ Transmissible change in germ cells
Translation
gets transmitted to all cells of the
● Assembling amino acids based on the
offspring
codes
● Mutations are ​rare
● mRNA is translated in codons (three
nucleotides)
Mutations
● Are random and rare
● Can be beneficial, neutral, or harmful for
the organism
● Do not try to supply what the organism
needs
● Random - whether a particular mutation
happens or not is unrelated to how useful
the mutation would be
● Not all mutations matter
● Somatic mutation
○ Occur in non-reproductive cells
○ Won’t be passed onto offsprings
● Germline mutations
○ Occur in sperms or ova
○ Will be passed onto offspring
○ A section of DNA is lost, or deleted
● Frame Shift
○ One base pair is deleted and DNA
is replicated or transcribed which
shifts the whole reading
(frameshift)
○ Causes codons to be parsed
incorrectly
Bacterial Gene Recombination
● Vertical Gene Transfer - occurs during
reproduction​ between generations of cells

Causes of Mutation
● DNA fails to copy accurately
○ Most of the mutation that
we think matter to evolution
● Horizontal Gene Transfer - transfer of
are “naturally-occurring”
genes ​between cells of the same
● External influences can create
generation
mutations
○ Can be caused by exposure
to specific
■ Chemical
■ Radiation
■ Environmental
exposure
■ Endogenous factors
Types of Mutation Genetic Recombination
● Substitution ● Exchange of genes between two DNA
○ Nucleotide base is substituted for molecules
another (ex. A to G) ○ Crossing over occurs when two
■ May result to: chromosomes break and rejoin
● Sickle cell anemia ○ Recipient chromosome contains
○ Change a codon to one that new DNA
encodes the amino acid and
causes no change in the protein Bacterial Recombination
(silent mutation)
○ Change amino acid coding codon
to a sing stop codon and cause an
incomplete
● Insertion
○ Extra base pairs are inserted into a
new place in the DNA
● Deletion
 Transduction of Bacteriophage

Plasmids
● Self -replicating gene containing circular
species of DNA
○ Independent of the bacterial
chromosome
● F-factor - conjugative plasmid that carries
Bacterial Transformation
genes for sex pili
○ For transfer of plasmid to another
cell
● Encode proteins that enhance
pathogenicity o​ f bacteria
○ E.coli pathogenic plasmids
■ Enhance toxin production
and bacterial attachment
■ Without plasmid, E. coli is
benign
● Dissimilation plasmids - encode enzymes
for catabolism of unusual compounds
● R-factors - encode antibiotic resistance
Plasmid: R-Factor
Conjugation
Transposons