The microbial cell Flashcards
Prokaryotic vs eukaryotic
Pro = free floating genetic material, mesosome (organelle of bacteria) invagination of the plasma membrane functions either in DNA replication and cell division or excretion of exoenzymes
Euk = true nucleus, membrane-bound organelles of bacterial ancestry (mitochondria, plastids)
Shapes of bacterial cells
Cocci = spherical-shaped, ovoid, rounded
Bacillus = rod-shaped
Spirillum = spiral-shaped
Archaea cells have 4 different types of shapes
Prokaryotic cell envelope
Structure – cell membrane, cell wall, outer membrane (only in gram-neg bacteria)
Function – maintains shape, provides protection, prevents protection, prevents bursting/rupture in hypotonic environment
1. Cell membrane
- Bacterial phospholipid bilayer = glycerol phosphate head attached to fatty acid
- Cell membrane is not enough protection for prokaryotes = lipid nature makes prokaryote cell membrane vulnerable to chemicals, most pro surround cell membrane with a tough armour (cell wall), in gram-neg bacteria have additional outer membrane
2. The cell wall
- Consists of murein = type of peptidoglycan (sugar polymer w side chains of amino acids)
- Glycan chains are made of alternative units of N-acetylglucosamine and N-acetyl muramic acid connected by a beta 1-4 glycosylic bond
- Glycan chains are crosslinked together via a small (4-5 amino acids long) peptide attached to each N-acetyl muramic acid unit
- Murein is what helps bacteria to maintain shape
3. Outer membrane
- Only gram-neg bacteria have = asymmetric bilayer with lipopolysaccharides outer later and phospholipid inner layer
- LPS is lipid modified with sugars
- Each LPS has one glycolipid (lipid A), polysaccharide core (LPS core), long carbohydrate chain (up to 40 repeated sugars (O antigen)) confers hydrophobicity to the membrane
- O antigen = very immunogenic, elicits a strong antigenic response in vertebrates, made up of varying numbers of repeating units (very long = restricting access to the bacteria surface), not all gram-neg bacteria have O antigen but even without outer membrane is an effective barrier but can make nutrient uptake difficult
- Some bacteria do not have an outer membrane made of LPS, instead have waxes called mycolic acids (60-90 carbon long fatty acids), mycolic acids orient themselves in lipid bilayers, porins embedded in mycolic acid bilayer, mycolic acid bilayers are attached to peptidoglycan cell wall via complex sugars (arabinogalactan)
Gram positive cells
Stains purple
Cell membrane is surrounded by thick cell wall made of peptidoglycan/murein
Teichoic acids gives cell wall rigidity and helps gram-pos bacteria adhere to surfaces
Cell wall not as secure a defence
Beta-lactams are a class of antibiotics that inhibit peptidoglycan synthesis:
1. Bind and inactivate the enzyme (penicillin binding protein (PBP)) that assembles peptidoglycan chains and bonds the peptides of the peptide glycan layers to each other
2. Without active enzyme cell wall cannot be remodelled as cell grows
3. Cell wall weakens so much that internal osmotic pressure eventually causes cell to burst
Gram negative cells
Stains red
Cell membrane surrounded by thin cell wall and outer membrane, which is chemically different from other membrane with an asymmetric bilayer and very resistant to harmful chemicals
1. Application of crystal violet
2. Application of iodine (mordant)
3. Alcohol wash (decolorisation)
4. Application of safranin (counterstain)
Porins
Protein channels in the membrane of gram-neg bacteria
Mediate the diffusion of hydrophilic compounds – sugars, amino acids, ions
Periplasm
Aqueous/gel-like space between cell membrane (inner) and outer membrane
Murein layer found within
Contains = degradative enzymes that break down molecules so they can be transported across the inner membrane, proteins with affinity for sugars and amino acids that equip the cell to soak nutrients from the growth medium, beta-lactamases which protect the cells by inactivating antibiotics
Acid fast bacteria
Bacteria with outer membrane made of mycolic acids
Mycolic acids made Myobacterium tuberculosis colonies look like yellowish lumps of wax
Myco = fungus because of the way the colonies grow
Because the outer membrane is very hydrophobic these bacteria are resistant to harsh chemicals and desiccation
Cannot be identified with gram stain
Acid fast stain: = acid fast bacteria remains red, non-acid fast bacteria turn clear then stain blue
1. Stain with carbol fuchsin (red) in the presence of heat (to loosen waxy layer). Any bacteria can be stained with it.
2. Wash with a mixture of acid and alcohol to decolorise all but acid fast cells.
3. Counterstain with methylene blue to show cells that are not acid-fast.
Mycoplasmas
Do not have cell wall, just cell membrane
Have sterols in cell membranes to make them more rigid and tougher
Mycoplasmas do not have peptidoglycan so beta-lactam antibiotics do not affect them
Additional exterior layers and appendages
Capsules and slime layers
- Many pro surround themselves with coat of slime
- Made up of high-molecular polysaccharides or polymers of amino acids
- When coat is attached to cell it is called capsule and when loose its called slime layer
- Only produced in response to certain environmental cues (coats only needed under particular conditions)
- Helps microbes retain water and nutrients and resist the diffusion of chemicals = e.g some make in response to antibiotics
- Helps microbes adhere to surfaces and build multicellular communities = biofilms
- Line of defense = microbes avoid being eaten up by wbc
Flagella
- Propel many pro through lipids and wet surfaces
- Mobility important for virulence and biofilm formation
- Have = long helical filament, connecting hook, basal body (equipped with rotor to turn flagellum)
- Use ATP to fuel
- Filament = helical, rigid, hollow, composed of single protein flagellin which is very antigenic
- Hook = joint between filament and basal body, made of single protein hook protein
- Basal body = rod attached with rings which anchor the structure to the cell envelope while allowing rod to rotate, consists of different proteins
Pili
- Pilus = hair
- Allow cell to attach to hosts and surfaces, transfer proteins and nucleic acids and move
- Most commonly mediate attachment via adhesins to surfaces such as mucosa
- Helps cells to escape attack by wbc, resist engulfment phagocytosis
- Very antigenic
- Made of proteins called pilins
- Common in gram-neg bacteria, rare in gram-pos
- Pili twitching mobility = extend and retract by adding or removing a pilin to the base of the pilus (undergo processes of polymerisation and depolymerisation):
1. Tip of pilus touches a surface, the adhesins at the tip adhere the cell to the surface
2. Pilus depolymerises its base, shortening of pilus drags the cell forward
Prokaryotic intracellular structure
Prokaryotic cells have 2 main intracellular structures – nucleoid and cytoplasm
Prokaryotes major distinction is the absence of a true nucleus, prokaryotic DNA is not confined by a membrane bound structure, prokaryotes compact their DNA within the cytoplasm in a structure called the nucleoid, this allows prokaryotes to couple transcription and translation
DNA organisation in prokaryotes
DNA organised in chromosomes
Prokaryote chromosomes are circular in most cases
Most bacteria have a single chromosome
DNA needs to be compacted 1000fold to fit into the cell
DNA double helix in the circular bacteria chromosome is folded by DNA-binding proteins into supercoil loops which protrude from a denser core (chromosome packed like a bottle brush)
Supercoils can be:
- Negatively supercoiled – DNA twisted in the opposite direction to double helix
- Positively supercoiled – DNA twisted in the same direction to double helix – organisms in extreme environments
- Most bacterial genomes are negatively supercoiled
DNA replication in prokaryotes
DNA replication starts at the origin of replication (oriC) where a specific protein (DnaA) binds and separates the two DNA strands
Specific proteins are recruited at each of the two replication forks and DNA replication is carried out from the oriC in each direction
DNA replication stops when DNA replication machinery enters the terminus region (terC)
DNA replication in prokaryotes has 3 major challenges
1. Unwinding the DNA helix
2. Replicating each DNA strand simultaneously
3. Separating the two daughter chromosomes
Unwinding the double helix
After binding of DnaA has separated the two DNA strands:
- DnaB helicase is responsible for separating the two parental DNA strands
- The strands are kept apart by single stranded DNA binding proteins (SSB)
Separating and relacing the two strands of DNA in a circular DNA molecule tightens the double helix in the unopen region and induces the formation of positive supercoils, supercoils which can block DNA replication
To stop positive supercoiling, DNA gyrase eases the tension (positive supercoil) formed by the rapid unwinding of the helix carried out by the DnaB helicase
Replicating each DNA strand simultaneously
- DNA primase synthesises the RNA primer
- The leading strand is synthesised continuously
- The lagging strand is synthesised discontinuously and produces Okazaki fragments
- Okazaki fragments are ligated, and primers are removed
- Each new DNA molecule contains one template strand from the original molecule and one new DNA molecule
