Chapter 6: Bacterial Structure, Growth

1
Chapter 6 Bacterial Structure, Growth
Metabolism

• Chapter Objectives
• Be familiar w/ all aspects of Bacterial Cellular
  organization Structures, Functions and
  Compositions.
• What is Chemotaxis?
• Be able to fully distinguish Gram () cell walls
  from Gram (-) cell walls.
• What are Plasmids, Inclusion Bodies?
• Know the 2 major bacterial spore formers (Genus
  name).
• Be familiar with all of the environmental factors
  affecting bacterial cell growth.
• Know how bacteria reproduce be familiar w/ the
  4 major phases of bacterial growth.
• What is Metabolism?
• Know the 3 Biochemical Mechanisms utilized by
  bacteria for their E Production.
• Laboratory Be familiar w/ the steps comprising
  the Gram Stain Procedure.

2
Overview (Flashback)

• ALL bacteria are Prokaryotes.
• Prokaryotic cells lack a nucleus.
• Eukaryotic cells possess a well-defined nucleus
  (i.e., Human cells).
• Bacterial DNA is NOT organized into chromosomes
• Single double-stranded structure instead.
• Prokaryotes Eukaryotes w/ similar metabolic
  pathways cell growth, viability

3
Cellular Organization of Prokaryotic CellsNote
All bacterial cells possess a Cell Envelope,
Glycocalyx, Cell Membrane, Cell Pool, Ribosomes
and a Nucleoid the majority of bacterial cells
also contain a Cell Wall. Flagella, Fimbriae,
Pili, Capsules and Slime Layers all common to
many bacteria, but NOT universal to all bacteria.
(Fig. 6.1 p.49)
4
Appendages

• Structures for Motility Attachment (exterior)
• Flagella (singular, Flagellum) Motility
  Appendage
• Longest appendage
• Attached to Cell Wall, Cell Membrane or BOTH
• Via Basal Body complex molecular machine for
  rotation
• Function Directed movement influenced by () or
  (-) Chemotaxis (survival function) propel
  bacteria
• Composition
• Long, hair-like appendages composed of repeating
  units PRO Flagellin vary in and arrangement,
  usu. several 1000
• Long semi-rigid, helical, hollow tubular
  structure
• Highly Antigenic Flagella are Ags
  characteristic of bac.

5
Appendages Contd.

• Structures for Motility Attachment (exterior)
• Attachment Appendages
• Pili (sex pili)
• Fimbriae (adhesions, lectins, evasins or
  aggresins)
• Pili (singular, Pilus)
• Function
• Contributes to bacterial attachment and
  subsequent bacterial colonization infection
• Attachment organs, promoting specific
  cell-to-cell contact between Bac cells, or Bac
  cell Eukaryotic host cell
• Composition
• short, hair-like appendages composed of PRO Pilin
• Shorter and thinner than flagella
• Limited in number to 1 - 3 per cell

6
Appendanges Contd.

• Structures for Motility Attachment (exterior)
• Fimbriae (synonymous w/ Pili)
• Function
• Contributes to bacterial attachment and
  subsequent bacterial colonization infection
• Attachment organs, promoting specific
  cell-to-cell contact between Bac cells, or Bac
  cell Eukaryotic host cell
• Composition
• Small bristle-like fibers composed primarily of
  PRO Pilin
• Smaller in length and more numerous than pili
• s vary from a few to several 100 per cell
• Form a fringe on the bacterial cell surface

7
Appendages Contd.

• Structures for Motility Attachment (exterior)
• Sex Pilus (specialized Pilus)
• Conjugation or mating process
• Elongated, rigid, hollow tubular structure thru
  which DNA may be transferred
• Attachment between 1 bacterial cell and another
• Rate limiting factor of Sex Pilus Length
• Both Pili Fimbriae are Virulent features
• Contribute to the antigenicity of bacterial cells
• Note Bacterial Colonization precedes bacterial
  infection

8
Cell Envelope

• External (Outer) wrapping of Bacteria
• all material external to enclosing cytoplasm
• Glycocalyx secreted, forming EC coating
• Function
• Protection extreme environmental conditions
  loss of H2O (slows dehydration) and nutrients
• Attachment binds bac cells to other cells /or
  environmental surfaces.
• Structure
• Gelatinous polymer composed of ploysaccharides,
  proteins or BOTH
• Bacillus anthracis gt poly-D-glutamic acid
• Viscous (sticky) gelatinous consistency
• If tightly bound w/ organized structure Capsule
• Thick gelatinous polymer firmly attached to cell
  wall contributes to bac virulence by protecting
  bac from phagocytosis by WBCs that engulf
  destroy microbes.
• Increases bacteria survival direct contribution
  to pathogenicity
• If loosely bound amorphous Slime Layer
• Loose, soluble shield non-uniform in density
  protects cell from loss of H2O nutrients aids
  in attachment virulence capabilities

9
Cell Envelope Contd

• Cell Wall - Exoskeleton of the Bacterial Cell
• Function
• Gives/maintains SHAPE of bacterium high tensile
  strength!
• Protects cell from osmotic lysis
• Protects fragile cytoplasmic membrane
• Allows for influx of sm. molecules fluids d/t
  porosity
• Composition
• Peptidoglycan (PG) x-linked polymeric mesh (fig.
  6.2 p.50)
• Long polysaccharide chains of 2 repeating
  disaccharides (glycan chains) backbone of
  alternating glycan (monosaccharide) molecules
• N-acetylglucosamine (NAG)
• N-acetylmuramic acid (NAM)
• Glycan chains are connected by x-linker
  tetrapeptides or pentapeptides (Ex. Pentaglycine
  bridge), which connect NAM to each other and
  other peptides, resulting in interbridge
  formation.
• Short string of AAs gt serve to x-link adjacent
  polysaccharides at NAM subunits, forming network
  w/ high tensile strength.

10
Cell Envelope Contd.

• Cell Wall of Gram () Bacteria
• Composition
• Multiple layers of PG usu. 40 layers
• 90 of gram () cell wall PG
• Resists omotic lysis to a greater degree than
  gram (-) bacteria
• Acidic polysaccharides
• Teichoic acid polymer of glycerol units linked
  by phosphodiester bonds
• On exterior
• Major cell surface antigens
• Lipoteichoic acid
• Embedded w/in phospholipid bilayer of cell
• Function of Teichoic/Lipoteichoic acid
• Transport of ions
• Reinforces x-linking between glycan chains
• Enhances rigidity of cell wall

11
Cell Envelope Contd.

• Cell Wall of Gram (-) Bacteria
• Composition 3 layered cell wall
• Outer membrane largest part
• PG 1 single layer ONLY
• Periplasmic space
• An extensive region BETWEEN inner outer
  membrane
• Outer membrane
• Composition
• Phospholipid Bilayer (similar to inner memb)
• Specialized proteins polysccharides
  interspersed w/in bilayer
• Lipopolysaccharides
• Lipoproteins Lipid-pro complex anchors outer
  membr to PG layer (w/in Periplasmic space)
• Porin proteins protein channels completely
  span outer memb func transport small molecules
  across bilayer
• Location
• External to PG

12
Cell Envelope Contd.

• Cell Wall of Gram (-) Bacteria
• Outer Membrane
• Function
• Protective barrier
• Selective permeability
• Periplasmic Space
• Location
• region between inner outer membrane incl. PG
  layer
• Contents
• Degradative or Hydrolytic enzymes
• Phosphatases, nucleases, proteases lipases
• Transport proteins for sugars AAs

13
Cell Envelop Contd.

• Cell Membrane (Cytoplasmic Membrane, Protoplasmic
  Membrane, Inner Membrane)
• Composition
• Phospholipid Bilayer w/ peripheral integral
  proteins
• Peripheral Proteins do NOT cross entire bilayer
• Found on or extending to surface
• Integral Proteins embedded completely w/in
  entire bilayer
• Both Proteins aid in transport of solute
  molecules into the cell
• Phospholipid Molecule consisting of 3 units
• Polar Head Group gt Phosphate Group (Hydrophilic
  or H2O loving)
• 2 Non-polar Tails gt 2 long-chain FAs
  (Hydrophobic or H2O fearing)
• Hydrophilic phosphate groups point outward
• Non-polar lipid (hydrophobic) tails sequestered
  to interior

14
Cell Envelope Contd.

• Cell Membrane
• Location
• Very thin, flexible structure lying inside the
  cell wall, and molded completely around the
  cytoplasm.
• Function
• Retains cytoplasm protection for interior
  cellular structures
• Semi-permeable or selectively-permeable barrier
• Allows for transport of H2O, CO2, O2, simple
  sugars waste products
• Site of metabolic activities attributable to
  Eukaryotic organelles
• Respiration Photosynthesis gt synthetic rxns
  ATP production
• Transport systems
• Synthesis of cell wall components
• Synthesis of lipids
• Excretion of exoenzymes (essential for nutrient
  processing)
• Excretion of toxins (Clinical Note this aids in
  the disease process)

15
Summary (Fig. 6.3 p.51)Gram () Bacteria vs.
Gram (-) Bacteria

• Gram () Bacteria
• Thick, multilayered PG cell wall exterior to
  membrane
• PG covalently linked to Teichoic acid
• Gram (-) Bacteria
• 2 membranes
• Outer Membrane
• Inner (Cytoplasmic) Membrane
• PG layer is between inner outer membrane w/in
  the Periplasmic Space
• Periplasmic Space w/ enzymes and other proteins
• PG layer is THIN clinically relevant WHY?
• Outer membrane w/ lipopolysaccharides
• O-polysaccharide (polysaccharide portion)
  Antigenic
• Lipid-A (lipid portion) toxic integral memb.
  protein (ENDOTOXIN)

16
Protoplasm Internal Contents of Cells

• Protoplasm (Cytoplasm)
• Dense gelatinous soln.
• Primary component H2O (70-80)
• H2O solvent for the cell pool
• Cell Pool
• complex mixture of nutrients CHOs, PROs
  (enzymes), AAs, lipids inorganic salts (ions)
• Components of cell pool building blocks for cell
  synthesis or sources of energy
• Raw material for all metabolic rxns

17
Protoplasm Internal Contents of cells Contd.

• Mesosomes (Internal Membranes)
• Location
• Cell membrane extends inwardly into coiled
  passages or sacs
• Function
• Increases internal surface area for metabolic
  activities therefore site of many enzymes
• May be involved in secretory processes
• Septal Mesosomes located near site of cell
  division or septum wall formation anchoring
  sites for DNA molecule during chromosomal
  replication distribution to daughter cells.
• Bacterial Chromosome (chromatin or nuclear body)
• Single circle of dsDNA (limited amnt. of DNA
  needed for bac. cell survival)
• DNA is ed to irregularly-shaped area known as
  Nucleoid or Nuclear region
• Minimal genetic requirement for survival
• Genetic info. for cells structure function
• Not enclosed by a membrane
• Chromosome is attached to septal mesosome,
  usually.
• Located near site of cell division or septal wall
  formation

18
Protoplasm Internal Contents of cells Contd.

• Plasmids (extra DNA)
• Tiny, circular extrachromosomal strands of DNA
• Non-essential pieces of DNA
• Protective traits or selective advantages
• Antibiotic resistance
• Production of toxins
• Production of enzymes
• Important role in Genetic Engineering plasmids
  can be transferred from one bacterial cell to
  another
• Ribosomes
• Location
• Free in Cytoplasm or attached loosely to plasma
  membrane or mesosomes
• Composition
• Chemically combination of rRNA (60) protein
  (40)
• Bacterial ribosomes sediment _at_ 70S composed of 2
  subunits 50S and 30S
• Function
• Involved in protein synthesis
• When protein synthesis, found in chains of
  polyribosomes

19
Protoplasm Internal Contents of cells Contd.

• Inclusion Bodies/Granules
• Location
• w/in cytoplasm matrix
• No discernible structure to these globules
• May or may not be covered w/ membrane
• Function
• storage of food in times of plenty reserve
  deposits produced during nutrient abundance
• May vary in size, , content
• 2 Types
• Organic Inclusion Bodies
• Polysaccharide granules glycogen or starch
• Lipid granules polymer poly-ß-hdroxy-butyric
  acid (PHB)
• Inorganic Inclusion Bodies
• Polyphosphate granules inorganic phosphate
• Building blocks for nucleic acids ATP (E)
  production

20
Protoplasm Internal Contents of cells Contd.

• Endospores (spores)
• Formed w/in bacterial cell, in the cytoplasm
• Released from original cell as free spores (fig.
  6.5 p.53)
• Resistant or Dormant Structure
• Resistance to environmental stresses
• Heat, freezing
• UV radiation
• Chemical disinfectants
• Desiccation (drying)
• Dormant Structure dehydrated state of mature
  spore
• No metabolic activity
• No cell division
• Initiator of Spore Formation
• Unfavorable growth conditions nutrient depletion
• Note 1 vegetative cell or bacterial cell ? 1
  Spore

21
Protoplasm Internal Contents of cells Contd.

• Presence of Spores
• Genus Identification Species Identification
• -Clostridium -Location of
  Spore
• -Bacillus 1.
  Terminally
• 2.
  Sub-terminally
• Ex. Bacillus anthacis anthrax Bacillus cereus
  gastroenteritis Clostridium tetani tetanus
  Clostridium botulinum botulism Clostridium
  perfringens gas gangrene

22
Protoplasm Internal Contents of cells Contd.

• Endospores (spores)
• enhance survivability in hostile environments
• dormant cell within the bacterial cell
• Spores most resistant life forms
• Sporulation/Sporogenesis
• Process by which bacterial cell forms a spore
• Repackaging a copy of bac DNA into NEW form
• Decrd H2O, decrd metabolic activity, no
  division
• Restructured, impermeable, multilayered envelop
• Cell committed to this Sporangium (mother cell)

23
Protoplasm Internal Contents of cells Contd.

• Sporulation/Sporogenesis
• Begins w/ invagination of parent cell membrane
• Spore septum formation
• Septum forms a double layered membrane enclosing
  the protoplasm internal cellular components
  (incl. copy of bacterial DNA gt Forespore
  formation
• Protection to forespore
• Layer of PG
• Thick cortex
• Spore-coat keratin-like protein
• Impermeable to H2O thus maintenance of
  dehydrated state
• Mature spore then retains complete machinery for
  PRO synthesis new spore-specific enz. production
• Release of endospore by parent cell lysis

24
Protoplasm Internal Contents of cells Contd.

• Spore Germination
• Process by which the bacterial cell returns to
  vegetative state when favorable environmental
  conditions arise (germinating agents)
• Strong oxidizing agents
• Low pH
• Heat exposure
• Formation of hydrolytic (digestive) enzymes
• Degradation of spore coat, spore walls, cortex
• Germinating agents initiate activation of spore
• Presence of H2O nutrients enable outgrowth of
  spore
• Outgrowth dependent upon rehydration of spore w/
  incr. in biosynthetic activity

25
Bacterial Growth Metabolism

• All cells (bacterial human) undergo metabolic
  rxns to maintain life
• Environmental factors affecting bacterial cell
  growth
• Availability of Nutrients H2O
• Temperature
• Presence or absence of gases
• Osmotic pressure, esp. of external solute to
  bacterial cell
• pH

26
Bacterial Growth Metabolism

• Environmental factors affecting bacterial cell
  growth
• Nutrients
• Generation of energy
• Synthesis of cellular materials (metabolic rxns)
• Essential or Common nutrients (basic bioelements
  needed for bacterial cell growth)
• H2O 70-90 cellular wt universal solvent
  hydrolyzing agent
• Carbon food E source in form of PRO, Sugar,
  Lipid
• Nitrogen for PRO syn reqd in nucleic acid syn
  (purines pyrimidines)
• Sulfur (sulfate) AA syn (i.e., Cystine)
• Phosphorus (phosphate) provided by Pi key
  component of DNA RNA, ATP, and inner outer
  membrane phospholipids
• Minerals assocd w/ PRO (i.e., FePRO) common
  component of enzymes

27
Bacterial Growth Metabolism

• Environmental factors affecting bacterial cell
  growth
• Nutrients
• Essential or Common Nutrients 2 types
• Macronutrients needed in large quantities b/c
  reqd for cellular metabolism basic cell
  structure
• C, H, O, N
• Micronutrients needed in small/trace amnts
  more specialized func (enzyme pigment structure
  func)
• Trace elements (i.e., Mn Zn)
• Fastidious Bacteria microbes that require other
  complex nutrients/growth factors ( i.e., Vitamins
  or AAs)

28
Bacterial Growth Metabolism

• Environmental factors affecting bacterial cell
  growth
• Temperature
• Categorized optimum temp range w/ optimal temp
  (growth rate maximal)
• Min Max in each temp range determined by
  stability functionality of bac cell membranes
  and proteins.
• Max temp determined by PRO stability WHY?
• Min temp defined by PRO syn on ribosomes.
• 3 Categories for optimal bac growth (next slide)

29
Bacterial Growth Metabolism

• Environmental factors affecting bacterial cell
  growth
• Temperature
• 4 categories for bacterial cell growth
• Psychrophile (cold-lovers)
• Maximal growth _at_ 15C
• Range (-10 25)C
• Unable to cause humans dis b/c normal body temp
  37C
• Mesophiles (middle-lovers)
• Maximal growth _at_ 37C
• Range (20 40)C
• Primary grp of microorganisms causing human
  infection
• Thermophiles (heat-lovers)
• Maximal growth _at_ 65C
• Range (45 70)C
• Unable to cause human disease found in natural
  hot springs
• Hyperthermophiles range (70 110)C

30
Bacterial Growth Metabolism

• Environmental factors affecting bacterial cell
  growth
• Presence or Absence of Gases
• Primary gases O2, N2, CO2
• O2 w/ greatest impact on microbial growth (even
  if the microorganism does not require it)
• Oxygen Requirement for Bacterial growth
• Aerobic O2 present
• Obligate aerobe 20 O2 only grows w/ O2
• Microaerophile 4 O2 best growth w/ sm. amount
  O2
• Anaerobic O2 depleted
• Obligate anaerobe only grows in absence of O2
• Aerotolerant anaerobe anaerobes that tolerate
  /or survive in O2, but do NOT utilize O2 during
  E metabolism
• Facultative Anaerobe grows both in presence
  absence of O2 but grows BEST under Aerobic
  conditions considered to be aerobic organism O2
  present aerobic respiration for E O2 absent
  anaerobic pathways (fermentation)

31
Bacterial Growth Metabolism
32
Bacterial Growth Metabolism

• Environmental factors affecting bacterial cell
  growth
• Presence or Absence of Gases
• Growth of a microbe in Aerobic Environment
  (Aerobic microorganisms)
• Requirement for Catalase Superoxide Dismutase
• Function of enzymes break-down toxic
  intermediates of aerobic metabolism
• Catalase converts 2H2O2 ? 2H2O O2
• Superoxide Dismutase converts
• 2O2- 2H ? O2 H2O2

33
Bacterial Growth Metabolism

• Environmental factors affecting bacterial cell
  growth
• Osmotic Pressure or Osmolarity
• Tonicity of solute external to cell (
  soluteext )
• Osmosis diffusion of H2O thru a membrane
• Osmotic pressure considered wrt environment in
  which the cell resides.
• Relative tonicity of soln external to cell
• Isotonic soluteext cells soluteint
  osmosis proceeds _at_ same rate in both directions
  no net change.
• Hypotonic soluteext has lower that same
  solute inside cell, thus higher H2O outside
  cell net diffusion of H2O INTO the cell.
• Note bacteria in hypotonic environments do NOT
  burst swelling is restrained by presence of cell
  wall.
• Hypertonic soluteext is higher than
  soluteint, thus lower H2O outside the cell
  to achieve osmotic balance, there is an outward
  flux of H2O from cell to ext. environment
  osmotic outflow of H2O causes cell to shrink and
  become distorted.
• Osmophiles bacteria that reside in solns w/ high
  solute
• Halophiles (salt-lovers) osmophiles that
  require salt gt12

34
Bacterial Growth Metabolism

• Environmental factors affecting bacterial cell
  growth
• pH
• Majority of bacteria grow BEST _at_ neutral or
  slightly alkaline pH
• pH 7.0 7.4 gt this is near most normal body
  fluids
• Acidophiles grow BEST _at_ low pH (acid pH 0
  1.0)
• Alkalophiles grow BEST _at_ high pH (alkaline pH
  10.0)

35
Bacterial Growth Metabolism

• Bacterial Growth
• Bacteria reproduce Asexually by process known as
  Binary or Transverse Fission
• Binary 1 cell becomes 2
• Transverse the wall that completely crosses the
  parent cell splits the parent cell into 2
  daughter cells
• Preparation doubling of all cell components
  elongation of cell to 2x original size then it
  will completely cross divide the cell w/
  formation of transverse septum / transverse cell
  wall.

36
Bacterial Growth Metabolism

• Bacterial Growth
• Batch or Limited Growth Curve
• b/c bacteria reproduce by Binary Fission, the
  of cells incrs exponentially w/ time
  (exponential or log phase of growth)
• 1 cell becomes 2, 2 become 4, 4 become 8, 8
  become 16, etc.
• 1 batch just a limited supply of nutrients
  available
• System is closed
• No removal of waste products
• No monitoring of pH
• No aeration
• No exogenous nutritional supplements
• 4 major phases of growth (next slide)

37
Bacterial Growth Metabolism

• Bacterial Growth
• 4 major phases of growth (fig 6.6 p. 54)
• Lag Phase (NO growth)
• Period of cell adjustment / preparation
• Synthesis of components needed for subsequent
  growth
• Cell mass cell size incr. in readiness for
  division
• Growth rate 0 NO incr. in cell
• Log Phase (Balanced or Exponential Growth)
• Balanced growth all measurable components of
  the cell increase at the SAME rate (i.e., PRO,
  DNA, RNA, mass) doubling of cells
  (1?2?4?8?16?32, etc.)
• Exponential growth constant growth cells
  divide at constant rate
• Both cell mass cell incr. in coordinate
  manner
• Calculation of generation time or growth rate
  Nf(Ni)2n
• GROWTH RATE IS GREATEST
• GENERATION TIME IS SHORTEST
• Growth rate - of generations of new cell formed
  in one hour
• Generation time the time to go from one
  generation to the next

38
Bacterial Growth Metabolism

• Bacterial Growth
• 4 major phases of growth
• Stationary Phase
• Entry into stationary phase d/t
• Accumulation of waste products
• Exhaustion of nutrients
• Change in pH
• Decrease in O2 levels
• NO increase in of viable cells
• of viable cells of dying cells
• The overall total of living dead cells is
  HIGHER in stationary phase vs. log phase
• Most resistant to environmental changes (heat,
  cold, drying, radiation, etc)
• Stationary Phase w/ largest Total of cells
  (both living dead) Log phase w/ largest of
  Viable cells.

39
Bacterial Growth Metabolism

• Bacterial Growth
• 4 major Phases of Growth
• Decline or Death Phase
• Growth rate decreases
• Death rate increases
• Limited amount of cellular growth availability
  of nutrients exhausted
• Dead cells provide nutrients for remaining viable
  cells (thus, there are some living cells in this
  phase)
• Clinical Note Log Phase is MOST effective stage
  for human infection bacteria are growing the
  fastest generation time shortest

40
Bacterial Growth Metabolism

• Bacterial Metabolism E Production
• Metabolism sum total of all chemical rxns
  occurring w/in a cell or microorganism
• Metabolic Rxns
• Catabolic (Degradative) release energy d/t
  break-down of large molecules into smaller
  constituent molecules (bond breakage) are
  exergonic rxns.
• Anabolic (Synthetic) require energy for the
  synthesis of large cellular molecules
  structures from smaller constituent building
  block molecules (bond building) are endergonic
  rxns.
• 3 Biochemical Mechanisms Utilized (fig. 6.8 p.
  55)
• Aerobic Respiration
• Anaerobic Respiration
• Fermentation

41
Bacterial Growth Metabolism

• Bacterial Metabolism E Production
• 3 Biochemical Mechanisms Utilized
• Aerobic Respiration
• Molecular Oxygen (O2) serves as the final e-
  acceptor of the ETC
• O2 is reduced to H2O
• E-generating mode used by aerobic
  chemoheterotrophs
• General term applied to most human pathogens
• E source Oxidation of organic compounds
• Carbon Source Organic Carbon
• 3 Coupled Pathways Utilized
• Glycolysis
• Krebs Cycle or Tricarboxylic Acid Cycle or
  Citric Acid Cycle
• Respiratory Chain or Electron Transport Chain
  (ETC)

42
Glycolysis

• Carbohydrate Catabolism
• CHOs are highly reduced structures (thus,
  H-donors) excellent fuels
• Degradation of CHO thru series of oxidative rxns
• Products of Glycolysis 2 net ATP, 2NADH
  (reduced coenzyme carrier) and 2Pyruvate
  (metabolic end product)
• 2ATP are expended during Gylcolysis to intiate
  CHO catabolism

43
Krebs (TCA) Cycle

• Initial substrate modified end product of
  Glycolysis
• 2Pyruvate or 2Pyruvic Acid is modified to
  2Acetyl-CoA, which enters the TCA cycle
• Circuit of organic acids (TCA) series of
  oxidations and reductions (REDOX rxns)
• Eukaryotes Mitochondrial Matrix
• Prokaryotes Cytoplasm of bacteria Cell
  Membrane
• E in the form of electrons is transferred to e-
  carriers (NAD FAD)
• NAD FAD are reduced NADH and FADH2,
  respectively
• Products of Krebs Cycle 6NADH and 2FADH2, and
  2ATP

44
Bacterial Growth Metabolism

• Bacterial Metabolism E Production
• Anaerobic Respiration
• Utilizes same 3 coupled pathways as Aerobic
  Respiration
• Metabolic process in which inorganic compounds
  other than O2 serve as the final e- acceptor
• Ex. Nitrate (NO3), Sulfate (SO4), or Carbonate
  (CO3)
• Have higher E potential than O2 thus yielding
  lower of ATP
• Variable of ATP produced dependent upon final
  e- acceptor available
• Used as an alternative to aerobic respiration
• In Facultative organisms
• Is obligatory is some species
• In Obligate anaerobes

45
Bacterial Growth Metabolism

• Bacterial Metabolism E Production
• Fermentation
• Metabolic process by which an organic metabolic
  intermediate derived from a fermentable
  substrate serves as the final e- acceptor
• Technically, incomplete oxidation of Glucose
• End products are neither more reduced or oxidized
  than substrate thus NO REDOX exchange RXNs are
  possible, since catabolic rxn takes place at same
  E level
• ATP is produced in an endergonic (E-requiring)
  event
• Only utilizes glycolysis pathway
• Sugars are fermentable substrates b/c they yield
  reducible intermediates
• Ex. Glucose ? Lactic Acid intermediate pyruvic
  acid
• Glucose ? EtOH intermediate
  acetaldehyde

46
Bacterial Growth Metabolism

• Bacterial Metabolism E Production
• Fermentation
• Products are of great commercial value
• Alcoholic beverages (beer, wine, whiskey)
• Industrial solvents (butanol, acetone)
• Organic acids (lactic acid, acetic acid)
• 2 general Product Categories
• Alcoholic Fermentation in yeast species w/
  capacity to convert Pyruvic acid ? Ethyl alcohol
• Acidic Acid Fermentation
• Homolactic fermentation only produces lactic
  acid
• Heterolactic fermentation results in mix of end
  products lactic acid, acetic acid, or propionic
  acid
• Lactic acid production implicated in food
  spoilage (i.e., souring of milk)
• Benefits of lactic acid in food industry
  yogurt, sauerkraut, and pickled food processes
• Swiss cheese ex. of mixed acid fermentation rxn
  between lactic acid proprionic acid (for
  flavor) and CO2 production (resultant holes)

47
Chapter 7 Bacterial Genetics

• Chapter Objectives
• What is meant by the term genome?
• Be familiar with the bacterial genome.
• Be familiar with bacteriophages.
• Be able to define the 3 types of Gene Transfer as
  related to bacteria.
• Know what a Mutation is and be able to
  distinguish point from frameshift mutations.

48
The Bacterial Genome

• Genome the totality of a particular organisms
  genetic material.
• Chromosome
• Single, long piece of circular dsDNA
• Most bacteria w/ 2000-4000 genes
• Highly folded
• Plasmids
• Small, circular dsDNA
• Replicate independently of the chromosome
• Only 5-100 genes
• For toxins PRO that promote transfer of plasmid
  to other cells
• NO genes for essential growth replication
• 1-20 copies/cell
• w/ mobile DNA sequences Transposons that can
  move between plasmids between plasmid and
  chromosome repository for antibiotic resistance
  genes

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Bacteriophage (Phage)

• A virus that replicates w/in a bacterial cell
• Composition
• Single piece of nucleic acid encapsulated w/in a
  protective protein coat.
• Nucleic Acid either DNA of RNA (ds or ss)
• Variable size 3-200 genes
• Replicative Cylce (fig 7.2 p. 60) Note time
  lapse 30 min. (FAST!)
• Phage attachment to receptors on host cell
  surface
• Injection of Nucleic Acid into the host cell
• Protein coat remains behind
• Phage nucleic acid gets incorporated into host
  cells biosynthetic machinery, and takes it
  over!!
• Then phage can replicate its own genetic
  material, synthesize phage-specific proteins.
• Assembly of new, mature phage particles w/ DNA or
  RNA encapsulated by a phage coat.
• Release by host cell lysis result of lysozyme
  action on bac cell wall

50
Bacteriophage (Phage)

• Classification
• Virulent Phage
• Infection w/ death to host bacterium by lysis
• Release of many progeny phage particles
• 1 Phage particle can produce 100s of progeny
• Temperate Phage
• Infection w/ same fate as above (lysis)
• Alternate fate phage DNA can fuse/integrate w/
  host bacterium chromosomes
• Prophage state expression of phage genes
  repressed indefinitely by repressors encoded w/in
  phage genome.
• No new phage particles produced
• Host bacterium survives
• Phage DNA replicates along w/ host chromosome

51
Bacteriophage (Phage)

• Classification
• Lysogenic Bacteria
• Bacteria that carry a prophage gt Lysogeny
• Bacterial cell is therefore L y s o g i n i z e d
• Nonlysogenic bacteria become lysogenic w/
  infection by temperate phage
• Highly stable union (bacterium prophage)
• Destabilization from UV radiation
• Induction emergence of virus from latent
  prophage state HOW?
• If DNA damage
• Repression of phage genes abolished
• Prophage excised from host chromosome
• Replication of prophage
• Production of progeny phage particles
• Host cell lysis w/ subsequent viral release

52
Bacterial Gene Transfer

• 3 Mechanisms
• Conjugation
• Process by which bacteria transfer genes from one
  cell to another cell by direct cell-to-cell
  contact under the direction of plasmids
• Donor (male) recipient (female) connected via
  the sex pilus, thru which DNA can pass
• Contact results in a relatively stable cell pair
• Only known mechanism for natural transfer of
  genetic material between different bacterial grps.

53
Bacterial Gene Transfer

• 3 Mechanisms
• Transduction
• Transfer of genes from one cell to another via a
  phage vector (temperate phage) w/out cell-to-cell
  contact.
• 2 mechanisms
• Generalized Transduction random fragment of
  bacterial DNA (from phage-induced cleavage of
  bacterial chromosome) is encapsulated in a phage
  protein coat instead of phage DNA.
• Rare phage particle can infect another cell,
  injecting the bacterial DNA fragment into the
  cell
• Recipient cell is transduced if the fragment
  becomes integrated into its chromosome.

54
Bacterial Gene Transfer

• 3 Mechanisms
• Transduction
• 2 Mechanisms
• Specialized Transduction only those bacterial
  genes located on the bacterial chromosome in
  close proximity to the prophage insertion site of
  the transducing phage are transduced
• Phage acquires bacterial genes by excision from
  bacterial chromosome (abnormal rare)
• Transducing phage particle contains both phage
  and bacterial DNA

55
Bacterial Gene Transfer

• 3 Mechanisms
• Transformation
• Transfer of genes from one cell to another by
  means of naked DNA
• Only minor effect on gene flow in natural
  bacterial populations
• Useful experimentally for introducing a cloned
  gene into bacterial cells

56
Genetic Variation

• Mutation any change in the structure of genetic
  material, specifically in the DNA base sequence.
• May be result of either base-pair substitution or
  the insertion or deletion of a nucleotide.
• Unstable revert back to original state
• Some display no noticeable effect on the organism
• Stable cause some change in the characteristics
  of an organism

57
Genetic Variation

• Types of Mutations
• Base pair substitution or Point mutations
• M/C type of Mutation
• One purine is substituted for another, or one
  pyrimidine is substituted for another
• Transition Mutations
• If left uncorrected by DNA Polymerase
  proof-reading function, they may result in the
  incorporation of one different AA into the
  corresponding protein this may or may not
  adversely affect the phenotypic expression of the
  cell.

58
Genetic Variation

• Types of Mutations
• Frameshift Mutations
• 1 or 2 base pairs are inadvertently added to or
  deleted from the DNA code sequence.
• Occurs m/likely as a result of nick in on of the
  DNA strands, w/ resulting addition or deletion of
  bases.
• Resultant change or shift in the translation of
  all of the genetic message following the addition
  or deletion
• Ex. Correct Seq ACC / TAG / CTA / TCG But, a
  deletion of the 2nd Adenine would change the
  message to read ACC / TGC / TAT / CG.
• Ultimate result an entirely NEW seq of AA in
  corresponding protein the protein is usu.
  NON-functional, causing a pronounced phenotypic
  change or cell death.

59
Genetic Variation

• Mutagens
• A physical or chemical agent that increases the
  rate of mutation
• Physical agents
• X-rays
• UV light
• Chemical agents
• Nitrous acid
• Acridine dyes
• Alkylating agents
• Base analogs (nucleotide-type bases that are
  slightly different from the normal nucleotides in
  DNA)