Lecture 3 - Bacterial Cell Structure (PART 2/SlIDES 69 -115)

Other than peptidoglycan, what main material is woven into a Gram-positive cell wall?

Teichoic acids (polymers of glycerol or ribitol phosphate).

What three major roles do teichoic acids play in the cell envelope?

• Maintain structure and integrity of the cell envelope
• Sequester ions (trap and hold cations within the wall)
• Protect bacteria from harmful environmental substances

In Gram-positive bacteria, what acts as the major barrier to diffusion?

The plasma membrane (cytoplasmic/cell membrane).

How do host cells detect the intracellular presence of bacterial peptidoglycan?

NOD1 and NOD2 are intracellular pattern recognition receptors that detect bacterial peptidoglycan (PG) fragments.

Can peptidoglycan only be detected inside host cells?

No. While NOD1 and NOD2 operate intracellularly, other host receptor proteins detect peptidoglycan in extracellular spaces.

What biological responses occur once the host immune system detects bacterial peptidoglycan?

It stimulates immune responses, triggers inflammation, and regulates host-microbe interactions.

What is the overall structure of a Gram-negative cell envelope from the inside out?

1. Inner membrane (plasma membrane)
2. Periplasmic space: Contains enzymes and a thin layer of peptidoglycan (occupies 20–40% of cell volume)
3. Outer membrane (OM): Protective asymmetrical lipid bilayer sitting atop the periplasm

What makes the Gram-negative outer membrane unique compared to standard membranes?

It is asymmetrical; its outer leaflet is composed almost entirely of lipopolysaccharide (LPS) rather than simple phospholipids.

Which molecules anchor the Gram-negative outer membrane to the cell wall?

Braun's lipoproteins, which serve as structural bridges connecting the outer membrane to the peptidoglycan layer across the periplasmic space.

What are the three structural components of Lipopolysaccharide (LPS) from bottom to top?

1. Lipid A (bottom): Anchors LPS in the outer leaflet; disaccharide with 4–7 acyl chains; acts as endotoxin
2. Core oligosaccharide (middle): Conserved sugar structure directly attached to Lipid A
3. O side chain / O-antigen (top): Variable repeating polysaccharide chain extending outward into the environment

Why is Lipopolysaccharide (LPS) critical for Gram-negative bacteria?

• Permeability barrier: Blocks entry of antibiotics, bile salts, and toxic chemicals
• Structural stabilization: Imparts an overall negative charge and stabilizes the outer membrane
• Host defense evasion: Shields against host immune attacks
• Endotoxin activity: Its Lipid A triggers potent immune and inflammatory responses

How does molecular entry into a Gram-negative cell differ from a Gram-positive cell?

In Gram-negative cells, molecules must first pass through porins (protein channels) in the outer membrane via facilitated diffusion before crossing the periplasm and reaching the plasma membrane.

What three layers can exist outside the cell wall or outer membrane, and what are their primary purposes?

1. Capsule
2. Slime layer
3. S-layer

Primary purposes:

• Protection: Guards against desiccation, environmental stress, phagocytosis, and predators
• Adhesion: Mediates attachment to surfaces and host tissues
• Biofilm formation: Promotes aggregation into communal bacterial networks

S-layer: structure, location, and primary functions

Structure & Location:

• Crystalline, lattice-like 2D protein/glycoprotein layer
• Located outside the peptidoglycan (Gram-positives) or outer membrane (Gram-negatives)

Functions:

• Protects against pH and ion fluctuations, osmotic stress, and enzymes
• Aids in surface adhesion
• Contributes to biofilm formation

Bacterial capsules: structural characteristics and protective functions

Structure:

• Highly organized and structured layer, firmly attached to the cell
• Typically composed of polysaccharides covalently linked
• Resistant to being washed off

Functions:

• Prevents phagocytosis by host immune cells and protozoan predators
• Protects against desiccation
• Contributes to biofilm formation

Bacterial slime layers: structural characteristics and protective functions

Structure:

• Unorganized, loose, and diffuse layer
• Composed of polysaccharides loosely associated with the cell
• Easily washed away

Functions:

• Protects against desiccation
• Helps resist immune predation
• Aids in gliding motility and biofilm formation

What are the five major functions of external bacterial structures?

1. Protection against environmental and immune threats
2. Attachment to surfaces (adhesion)
3. Horizontal gene transfer (e.g., conjugation)
4. Cell movement (motility)
5. Cell-cell interactions with other microbes, hosts, or predators

Standard Fimbriae/Pili vs. Sex Pili: structural and functional comparison

Fimbriae / Standard Pili:

• Structure: Short, thin, hair-like appendages; ~1,000 per cell
• Function: Attachment to surfaces, twitching motility, and DNA uptake

Sex Pili:

• Structure: Longer, thicker tubular shafts; ~1–10 per cell
• Function: Required for conjugation (direct plasmid transfer); plasmid-encoded

What is the primary purpose of flagella, and what are their four main distribution patterns?

Purpose: Motility and surface attachment.

Distribution patterns:

• Monotrichous: Single flagellum at one pole
• Amphitrichous: One flagellum at each opposite pole
• Lophotrichous: Tuft/cluster of flagella at one or both poles
• Peritrichous: Flagella distributed uniformly over the entire cell surface

What are the three main structural parts of a bacterial flagellum?

1. Basal body: Embedded in the cell envelope; functions as the rotary motor
2. Hook: Short, curved tubular segment connecting the motor to the filament
3. Filament: Long, rigid helical hollow cylinder extending into the extracellular environment

How is the bacterial flagellar filament assembled?

Assembled from the inside out (flagellin subunits travel through the hollow filament center and polymerize at the tip).

How do bacterial flagella move, and how can they be visualized under microscopy?

• Movement: They spin and rotate like propellers driven by a basal motor
• Visualization: Too thin for standard bright-field microscopy; requires special staining to build up filament diameter

What are the major categories and types of bacterial motility?

• Flagellar motility: Swimming (liquid), Swarming (surfaces), Spirochete motility (internal)
• Non-flagellar motility: Twitching (Type IV pili), Gliding (surface tracking/slime)
• Directed movement: Chemotaxis (movement oriented toward or away from chemical gradients)

How does bacterial swimming motility operate (Run vs. Tumble)?

• Counterclockwise (CCW) rotation: Bundles flagella together to drive smooth, directional forward movement (run)
• Clockwise (CW) rotation: Unbundles flagella, causing the cell to stop and reorient randomly (tumble)

Flagellar motor: what are the roles of the rotor and stator?

• Rotor (moving component): Includes the C-ring (FliG protein); rotates and transmits torque to the shaft
• Stator (stationary component): Composed of MotA and MotB proteins; forms transmembrane proton-conducting channels

What energy source powers flagellar movement, and how is torque generated?

• Energy source: Proton Motive Force (PMF)
• Mechanism: Protons pass through MotA/MotB stator channels, releasing energy that generates torque to spin the rotor

What is swarming motility, and how do bacteria coordinate this movement?

• Definition: Coordinated, multicellular group movement across moist, solid surfaces
• Flagella type: Typically powered by numerous peritrichous flagella
• Mechanism: Cells secrete biosurfactants to lower surface tension, enabling collective sliding

How do spirochetes achieve motility using internal flagella?

• Axial fibril (endoflagella): Multiple flagella remain within the periplasmic space, wrapping around the cell cylinder
• Corkscrew motion: Rotation of these internal flagella flexes the outer membrane, propelling the helical cell forward like a corkscrew through viscous fluids

How do twitching and gliding motility function on solid surfaces without flagella?

• Twitching motility: Short, jerky movements mediated by the extension, tethering, and retraction of Type IV pili
• Gliding motility: Smooth, continuous movement along surfaces without visible appendages, often accompanied by polysaccharide slime secretion

What is chemotaxis, and how do bacteria navigate chemical gradients?

• Definition: Directed movement toward chemical attractants or away from chemical repellents
• Sensing: Transmembrane chemoreceptors measure temporal changes in chemical concentration
• Mechanism: When approaching an attractant, bacteria tumble less frequently, extending the duration of forward runs

What are the six major specialized structures found in the bacterial cytoplasm?

1. Cytoskeleton
2. Inclusions (nutrient storage reserves)
3. Microcompartments
4. Gas vesicles
5. Magnetosomes
6. Endospores

Bacterial cytoskeleton: definition and three primary functions

Definition: Network of polymerizing protein filaments homologous to eukaryotic actin, tubulin, and intermediate filaments.

Three primary functions:

1. Regulate and drive cell division
2. Direct intracellular protein localization
3. Determine and maintain cell shape

FtsZ: cellular role and mechanism in bacterial cell division

• Homology: Eukaryotic tubulin homolog found in most bacteria
• Mechanism: Polymerizes at mid-cell to construct the Z-ring
• Function: Constricts the dividing cell and coordinates synthesis of the division septum

Roles of MreB and CreS (crescentin) in bacterial cell shape

• MreB (actin homolog): Found in rod-shaped bacteria; organizes the peptidoglycan synthesis machinery to maintain rod morphology
• CreS (crescentin) (intermediate filament homolog): Localizes to one side of the cell to produce a curved/comma morphology

What are bacterial ribosomes, and what is their total sedimentation coefficient?

• Function: Ribonucleoprotein complexes that carry out translation (protein synthesis)
• Sedimentation size: 70S (composed of a small and large subunit)

What are the two subunits of a bacterial 70S ribosome and their associated rRNAs?

• Small subunit (30S): Contains 16S rRNA and ~21 ribosomal proteins
• Large subunit (50S): Contains 23S rRNA, 5S rRNA, and ~31 ribosomal proteins

What are bacterial microcompartments, and how do carboxysomes function?

Microcompartments: Protein-shelled complexes that sequester specific metabolic enzymes without a lipid bilayer.

Carboxysome mechanism:

1. Carbonic anhydrase converts bicarbonate into high local concentrations of $\text{CO}_2$
2. RuBisCO utilizes the trapped $\text{CO}_2$ to synthesize sugars, minimizing oxygenase side reactions

What are gas vesicles, and what physiological advantage do they provide to aquatic bacteria?

• Structure: Rigid, hollow protein cylinders permeable to gas but impermeable to liquid water
• Function: Provide buoyancy
• Purpose: Allow aquatic microbes to position themselves at optimal water depths for light and nutrients

What are magnetosomes composed of, and how are they organized within the cell?

• Composition: Intracellular crystals of magnetic minerals (magnetite $\text{Fe}_3\text{O}_4$ or greigite $\text{Fe}_3\text{S}_4$) enclosed in membrane invaginations
• Assembly: Formed via guided biological mineralization of iron
• Organization: Aligned in straight linear chains held by the actin-like protein filament MamK

How do magnetosomes assist aquatic bacteria in locating their optimal niche?

• Geomagnetic alignment: Crystals orient the cell along Earth's magnetic field lines, directing swimming along a vertical axis
• Synergy with aerotaxis: Combines with oxygen sensing to guide bacteria to the oxic-anoxic transition zone (OATZ) where microaerophilic nutrient conditions are ideal

What is a bacterial endospore, and what harsh environmental conditions can it resist?

Definition: A dormant, highly differentiated survival structure produced by select bacteria during starvation.

Resistance capabilities:

• Extreme heat and freezing
• Severe desiccation
• Ionizing and UV radiation
• Chemical disinfectants and enzymatic degradation
• Prolonged nutrient depletion

How long can endospores persist, and what triggers their reactivation?

• Persistence: Capable of surviving in a metabolic stasis state for centuries to millions of years
• Reactivation trigger: Senses favorable environmental conditions and nutrient availability via germinant receptors

What is sporulation, and how does the process proceed inside the mother cell?

• Definition: The developmental process of forming a resilient endospore
• Trigger: Starvation or severe nutrient limitation
• Duration: Takes approximately 6–10 hours
• Mechanism: Initiates with asymmetric cell division; the mother cell engulfs the forespore, synthesizes protective layers, and subsequently lyses to liberate the spore

What are the four primary locations of endospores within the bacterial cell?

1. Central: In the middle of the cell
2. Subterminal: Positioned between the center and one pole
3. Terminal: Positioned at the very pole/tip
4. Swollen sporangium: Terminal or subterminal spore causing prominent localized cell distension/bulging

What famous scientific finding demonstrated the extreme longevity of endospores?

Viable halotolerant bacteria were revived from endospores preserved within an intact 250 million-year-old salt crystal.

What is a vegetative bacterial cell, and when does an endospore revert to one?

• Vegetative cell: A metabolically active, growing, and replicating bacterial cell
• Reversion trigger: Initiated when an endospore encounters favorable environmental conditions and sufficient nutrients

What are the three sequential stages in the transition from an endospore to a vegetative cell?

1. Activation: Prepares the spore casing (often prompted by sublethal heating)
2. Germination: Triggered when germinant receptors bind nutrients (sugars/amino acids), initiating rehydration and coat degradation
3. Outgrowth: Protoplast swells, synthesizes new cell components, and emerges from the cracked spore casing to resume active growth