PROKARYOTES

In week 3, I continued with the prokaryote topic in Microbiology's class. This week, I learned about the components of prokaryotic cell:

1) Structures external to the cell wall

2) Cell wall                                        

3) Structures internal to the cell wall

Morphology of bacterial cell

STRUCTURES EXTERNAL TO THE CELL WALL

1. Glycocalyx
2. Flagella
3. Axials Filament
4. Fimbriae and Pili

1. Glycocalyx

The glycocalyx (capsule, slime layer, or extra cellular polysaccharide) is a gelatinous polysaccharide and/or polypeptide covering. The exact chemical composition varies depending on the species.

• Capsules are organized and firmly attached to the cell wall.
• Capsules may protect pathogens from phagocytosis.
• Capsules enable adherence to surfaces, prevent desiccation, and may provide nutrients.
• Slime layers are unorganized and loosely attached to the cell wall.
• Extracellular polymeric substance (EPS) is the description of a glycocalyx that is a component of a biofilm.

2. Flagella

Flagella are relatively long filamentous appendages consisting of a filament, hook, and basal body. Prokaryotic flagella rotate to push the cell. Motile bacteria exhibit taxis; positive taxis is movement toward an attractant, and negative taxis is movement away from a repellent.

Arrangment of Bacterial flagella

The structure of a prokaryotic flagellum

Flagella are anchored by pairs of rings associated with the plasma membrane and cell wall. Gram positive bacteria have only the inner pair of rings. The filament is composed of the globular protein flagellin, which is arranged in several intertwined chains that form a helix around a hollow core.

• Flagellin can vary in structure and is used to identify some pathogenic bacteria serologically. The flagellar antigens are referred to as H antigens.
• E. coli may express any of at least 50 different variants; serovars (serological variants) identified as O157:H7 are associated with food borne epidemics (O antigens are somatic antigens and are lipopolysaccharide complexes associated with the cell wall).

Flagella and Bacterial motility

3) Axial Filament

Spiral cells that move by means of an axial filament (endoflagellum) are called spirochetes. Axial filaments are similar to flagella, except that they wrap around the cell.

4) Fimbriae and Pili

Fimbriae and pili are short, thin appendages. Cells may have many fimbriae, which help the cells adhere to surfaces. Cells that have pili have only one or two.

• Pili join cells either for the transfer of DNA from one cell to another (sex pili) or are used for special types of movement; twitching, seen inPseudomonas aeurginosa, Neisseria gonorrhoeae and some strains of E. coli, or the gliding motility of myxobacteria.

Fimbriae

CELL WALL

The cell wall surrounds the plasma membrane and protects the cell from changes in water pressure.

The bacterial cell wall consists of peptidoglycan (or murein), a polymer consisting of NAG and NAM and short chains of amino acids.

Penicillin interferes with peptidoglycan synthesis.

The Gram-positive Cell Wall

Gram-positive cell walls consist of many layers of peptidoglycan and also contain teichoic acids. Teichoic acids may:

• bind and regulate movement of cations into and out of the cell
• prevent extensive wall breakdown and possible cell lysis during cell growth
• provide much of the cell wall's antigenicity

The Gram-negative Cell Wall

Gram-negative bacteria have a lipopolysaccharide-lipoprotein-phospholipid outer membrane surrounding a thin (sometimes a single) peptidoglycan layer. Gram-negative cell walls have no teichoic acids. The outer membrane protects the cell from phagocytosis and from penicillin, lysozyme, and other chemicals. Porins are proteins that permit small molecules to pass through the outer membrane; specific channel proteins allow other molecules to move through the outer membrane. The lipopolysaccharide component of the outer membrane consists of sugars (O polysaccharides) that function as antigens and lipid A, which is an endotoxin. Endotoxin causes fever and shock.

Gram-positive cell wall vs Gram negative cell wall

The gram stain mechanism

The cells are first stained with the primary stain, crystal violet. After about 1 minute of staining excess primary stain is washed off and the mordant, Gram's iodine, is applied for another minute.

• The iodine forms a complex with the crystal violet and the crystal violet-iodine complex becomes "trapped" inside the peptidoglycan.
• At this point cells with either type of cell wall appear purple, due to the presence of the crystal violet-iodine complex.

The next step is to decolorize (using acetone-alcohol) the cells for a period of time long enough to dissolve the outer membrane of Gram-negative cells and pull the crystal violet-iodine complex through the thin layer of peptidoglycan.

• If done properly, Gram-positive cells retain the crystal violet-iodine complex and appear purple while Gram-negative cells are decolorized (and appear colorless, no?)
• Note: decolorizing too long will result in both Gram-positive and Gram-negative cells appearing colorless, while decolorizing for too short a time will result in both cell types retaining the crystal violet-iodine complex and appearing purple.

After decolorization Gram-positive cells (purple) can be differentiated from Gram-negative cells (colorless) but since colorless cells are hard to see the final step in the Gram staining process is to use a counter-stain, safranin, to stain the Gram-negative cells pink.

• Gram-positive cells will also stain with safranin but it will not be seen on top of the purple crystal violet-iodine remaining in the cells.

Atypical cell wall

• Mycoplasma is a bacterial genus that naturally lacks cell walls; the presence of sterols in the plasma membrane protects from osmotic lysis.
• Mycobacterium is a genus that has mycolic acids in its cell walls, giving it a "waxy" cell wall that is resistant to decolorization with acid-alcohol when stained with carbolfuschin (and so is designated "acid-fast").
• Archaea have pseudomurein; they lack peptidoglycan.

Damage to the cell wall

• In the presence of lysozyme, gram-positive cell walls are destroyed, and the remaining cellular contents are referred to as a protoplast.
• In the presence of lysozyme (after disruption of the outer membrane), gram-negative cell walls are not completely destroyed, and the remaining cellular contents are referred to as spheroplasts.
• Protoplasts and spheroplast are subject to osmotic lysis.
• Proteus and some other genera can lose their cell walls spontaneously or in response to penicillin and swell into L forms (Lister Institute). L forms can live and divide and/or return to the normal walled state.
• Antibiotics such as penicillin interfere with cell wall (peptidoglycan) synthesis.

STRUCTURES INTERNAL TO THE CELL WALL

1. Plasma (cytoplasmic) membrane
2. Cytoplasm
3. Nuclear Area
4. Ribosomes
5. Inclusions

1) Plama (cytoplasmic) Membrane

• The plasma membrane encloses the cytoplasm and is a phospholipid bilayer with peripheral and integral proteins (the fluid mosaic model).
• The plasma membrane is selectively permeable.
• Plasma membranes carry enzymes for metabolic reactions, such as nutrient breakdown, energy production, DNA replication and photosynthesis in prokaryotes. 
• These reactions take place in mitochondria, the nucleus, and chloroplasts in eukaryotic cells.

Movement of materials across membrane

• Movement across the membrane may be by passive processes, in which materials move from areas of higher to lower concentration, and no energy is expended by the cell.
• In simple diffusion, molecules and ions move until equilibrium is reached.
• In facilitated diffusion, substances are transported by transporter proteins across membranes from areas of high to low concentration.
• Osmosis is the movement of water from areas of high to low concentration across a selectively semi permeable membrane until equilibrium is reached.

Osmosis

• In active transport, materials move from areas of low to high concentration by transporter proteins, and the cell must expend energy.
• In group translocation, energy is expended to modify chemicals and transport them across the membrane. 
• Once inside the cell the chemical modification prevents the transported substance from moving out of the cell. Example: glucose is converted to glucose-6- phosphate as it passes into the cell. 
• This traps it and allows the movement of more glucose into the cell even when extracellular concentrations may be low.

2) Cytoplasm

• Cytoplasm is the fluid component inside the plasma membrane.
• The cytoplasm is mostly water, which inorganic and organic molecules, DNA, ribosomes, and inclusions.

3) Nuclear Area

• The nuclear area contains the DNA of the bacterial chromosome.
• Bacteria can also contain plasmids, which are circular, extra-chromosomal DNA molecules.

4) Ribosomes

• The cytoplasm of a prokaryote contains numerous 70s ribosomes; ribosomes consist of rRNA and protein.
• Protein synthesis occurs at ribosomes; it can be inhibited by certain antibiotics. The difference between prokaryotic (70s) and eukaryotic (80s) ribosomes allows antibiotics to selectively target the prokaryotic ribosomes while sparing eukaryotic ribosomes.

5) Inclusions

Inclusions are reserve deposits found in prokaryotic and eukaryotic cells. Among the inclusions found in bacteria:

1. metachromatic granules (inorganic phosphate)
2. polysaccharide granules (usually glycogen or starch)
3. lipid inclusions
4. sulfur granules
5. carboxysomes (ribulose 1,5-diphosphate carboxylase)
6. magnetosomes (Fe3O4)
7. gas vacuoles.

Magnetosomes

• present in several gram-negative bacteria (Aquaspirillum magnetotacticum for example)
• act like magnets
• may be used to move downward until they reach a suitable attachment site
• have been demonstrated to decompose hydrogen peroxide in vitro so it has been suggested that magnetosomes may protect cells from hydrogen peroxide accumulation

ENDOSPORES

• Endospores are resting structures formed by some bacteria for survival during adverse environmental conditions.
• The process of endospore formation is called sporulation; the return of an endospore to its vegetative state is called germination.  Two genera that commonly form endospores are Bacillus and Clostridium.