Chapter #3: Bacteria and Archaea Flashcards Preview

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1

S/V Ratio

Many years it was thought that microbes were small because it increases the surface area to volume ratio. As this ratio increases, nutrient uptake and diffusion of molecules within the cell become more efficient, which in turn facilitates a rapid growth rate. Shape affects the S/V ratio. A rod with the same volume as a coccus has a higher S/V ratio than does the coccus. This means that a rod can have greater nutrient flux across its plasma membrane.

For bacteria to be very large, they must have other characteristics that max their S/V ratio, or their size must be of some selective value.

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Plasma Membrane

Selectively permeable barrier, mechanical boundary of cell, nutrient and waste transport, location of many metabolic processes (respiration, photosynthesis), detection of environmental cues for chemotaxis.

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Ribosomes

Protein synthesis. Cytoplasm of bacterial and archaeal cells often packed with them, and other ribosomes may be loosely attached to plasma membrane. Cytoplasmic ribosomes synthesize proteins destined to remain in cell, whereas plasma membrane ribosomes make proteins that will reside in the cell envelope or are transported to the outside.

Bacterial and archaeal ribosomes are called 70S ribosomes (80S in eukaryotes) and are constructed of a 50S and a 30S subunit. The S stands for Svedberg unit.

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Nucleoid

Localization of genetic material.

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Cell Wall

Provides shape and protection from osmotic stress, toxin substances. And in pathogens, it can contribute to pathogenicity.

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Flagella

Swimming motility.

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Endospore

Survival under harsh environmental conditions; only observed in Bacteria.

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Cell Envelope

The plasma membrane and all the surrounding layers external to it. The cell envelopes of many bacteria consist of the plasma membrane, cell wall, and at least one additional layer.

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Intracytoplasmic Membrane Systems

In addition to the plasma membrane, some bacteria have extensive intracytoplasmic membrane systems. These internal membranes and the plasma membrane have a common, basic design. However, they can differ significantly in terms of the lipids and proteins they contain.

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Fluid Mosaic Model

Proposes that membranes are lipid bilayers within which proteins float.

Within the lipid bilayer, small globular particles are visible; these have been suggested to be membrane proteins lying within the membrane lipid bilayer.

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Amphipathic

Being structurally asymmetric, with polar and nonpolar ends. Most membrane-associated lipids are amphiphatic.

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Peripheral Proteins

A type of membrane protein. Loosely connected to the membrane and can be easily removed. They are soluble in aqueous solutions and make up about 20-30% of total membrane protein.

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Integral Proteins

Type of membrane protein. Not easily extracted from membranes and are insoluble in aqueous solutions when freed from lipids.

They are amphiphatic, their hydrophobic regions are buried in the lipid while the hydrophilic portions project from the membrane surface. Carbohydrates often are attached to the outer surface of plasma membrane proteins.

The integral proteins carry out some of the most important functions of the membrane. Many are transport proteins used to move materials in and out of cell. Some part of protein secretion systems. Other involved in energy conserving processes.

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Bacterial Lipids

The plasma membrane is very dynamic. The lipid composition varies with environmental temperature in such a way that the membrane remains fluid during growth. For example, bacteria growing at lower temperatures have more unsaturated fatty acids in their membrane phospholipids-that is, there are one or more covalent bonds in the long hydrocarbon chain. At higher temperatures, there phopspholipids have more saturated fatty acids- those in which the carbon atoms are connected only with single covalent bonds.

Other factors also affect the lipid composition of membranes. For instance, some pathogens change the lipids in their plasma membranes in order to protect themselves from antimicrobial peptides produced by the immune system.

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Hopanoids

Bacterial membranes usually differ from eukaryotic membranes in lacking sterols (steroid-containing lipids) such as cholesterol. However, many bacterial membranes contain sterol-like molecules called hopanoids.

They are synthesized from the same precursors as steroids, and like the sterols in eukaryotic membranes, they probably stabilize the membrane. Hopanoids are also of interest to ecologists and geologists: the total mass of hopanoids in sediments is estimated to be around 10^11-12 tons-about as much as the total mass of organic carbon in all living organisms (10^12 tons)- and evidence exists that hopanoids have contributed significantly to the formation of petroleum.

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Periplasmic Space

One important feature of the cell envelope of gram-negative bacteria is a space that is frequently seen between the plasma membrane and the outer membrane in electron micrographs. It also is sometimes observed between the plasma membrane and the wall in gram-positive bacteria. This space is called the periplasmic space.

The subtance that occupies it is the periplasm. The nature of the periplasmic space and periplasm differs in gram+ and gram- bacteria.

In gram-negative bacteria, contains hydrolytic enzymes and binding proteins for nutrient processing and uptake; in gram-positive bacteria and archaeal cells, may be smaller of absent.

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Bacteria Cell Wall

The game-positive cell wall consists of a single, 20-80 nm thick, homogenous layer of peptidoglycan (murein) lying outside the plasma membrane.

In contrast, the gram-negative cell wall is quite complex. It has a 2-7-nm thick peptidoglycan layer covered by a 7-8 nm thick outer membrane. THe walls of gram-positive cells are more resistant to osmotic pressure because of the thicker peptidoglycan layer.

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Peptidoglycan Structure

Enormous, meshlike polymer composed of many identical subunits. Each subunit contains two sugar derivatives, N-acetyl-glucosamine (NAG) and N-acetylmuramic acid (NAM), and several different amino acids.

The amino acids form a short peptide consisting of four alternating D- and L-amino acids; the petide is conencted to the carboxyl group of NAM. The presence of D-amino acids protects against degradation by most peptidases, which recognize only the L-isomers of amino acid residues.

The meshlike peptidoglycan polymer is formed by linking the petidoglycan subunits together to form a peptidoglycan strand. These strands are then cross-linked to each other. The backbone of a peptidoglycan strand is thus composed of alternating NAG and NAM residues. The peptidoglycan strands are joined by cross-links between the peptides of adjacent strands.Each peptidoglycan strand is helical, and the peptides extend out from the backbone at right angles to each other. Thus each strand can become crosslinked to strands at each side, above, and below.

Many bacteria cross-link peptidoglycan by connecting the carboxyl group of the terminal D-alanine (position 4) directly to the amino group of diaminopimelic acid (position 3). Some other bacteria use a peptide interbridge instead. With or without a peptide interbridge, cross-linking results in an enormous peptidoglycan sac that's actually one dense, interconnected network. These sacs have been isolated from gram+ bacteria and are strong enough to retain their shape and integrity, yet they're relatively porous and elastic.

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Gram+ Cell Walls

Gram+ bacteria belong to the phyla Firmicutes and Actinobacteria. Most of these bacteria have thick cell walls composed primarily of peptidoglycan. In addition, their cell walls usually contain large amounts of secondary cell wall polymers, including teichoic acids.

The periplasmic space of gram+ bacteria lies between plasma membrane and cell wall, and is smaller compared to gram- bacteria. The periplasm has few proteins; probably because the peptidoglycan sac is porous and any proteins secreted by the cell usually pass through it.

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Teichoic Acids

Polymers of glycerol or ribitol joined by phosphate groups. Amino acids such as D-alanine or sugars such as glucose are attached to the glycerol and ribitol groups. The teichoic acids are covalently connected to peptidoglycan or to plasma membrane lipids; in the latter case, they are called lipoteichoic acids. Teichoic acids appear to extend to the surface of the peptidoglycan. Because they're negatively charged, they help give the gram+ cell wall its negative charge. Teichoic acids aren't present in gram- bacteria.

Function not clear, but many theories. One is to help create and maintain the structure of the cell envelope and to protect the cell from harmful substances in the environment. Some of them may be involved in binding pathogenic species to host tissues, thus initiating the infectious disease process.

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Exoenzymes

Enzymes secreted by gram+ bacteria.

Exoenzymes often serve to degrade polymeric nutrients that would otherwise be too large for transport across the plasma membrane. Those proteins that remain in the periplasmic space are usually attached to the plasma membrane.

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Gram+ Cell Walls: Proteins on Surface

Staphylococci and most other gram+ bacteria have a layer of proteins on the surface of the peptidoglycan. These proteins are involved in interactions of the cell with its environment.

Some are noncovalently attached by binding to teichoic acids or other cell wall polymers. For example, S-layer proteins bind noncovalently to polymers scattered throughout the cell wall. Enzymes involved in peptidoglycan synthesis and turnover also seem to interact noncovalently with the cell wall.

Other surface proteins are covalently attached to the peptidoglycan. Many covalently attached proteins have roles in virulence. For example, the M protein of pathogenic streptococci aids in adhesion to host tissues and interferes with host defenses In staphylococci, these surface proteins are covalently joined to the pentaglycine interbridge of the peptidoglycan.

An enzymes called sortase catalyzes the attachment of these surface proteins to the peptidoglycan. Sortases are attached to the plasma membrane of the cell.

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Gram+ Cell Walls: Acid-Fast Bacteria

The acid-fast bacteria, a taxon within the phylum Actinobacteria, have somewhat different cell wall structure than the typical gram+ wall just described. These bacteria include members of the genus Mycobacterium. Their cell walls are unique among the gram+ bacteria because they consist of other layers in addition to peptidoglycan. An important component of their walls is a group of fatty acids called mycolic acids. Some have proposed that the mycolic acids form a bilayer structure that's analogous to the outer membrane of gram- bacteria.

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Gram- Cell Walls

Much more complex than gram+ walls. The thin peptidoglycan layer next to the plasma membrane and bounded on either side by the periplasmic space usually constitutes only 5-10% of the wall weight.

The periplasmic space of gram- bacteria is also strikingly different from that of gram+ bacteria. It ranges in width from 1-71 nm. May constitute about 20-40% of total cell volume. When cell walls are disrupted carefully or removed without disturbing the underlying plasma membrane, periplasmic enzymes and other proteins are released and may be easily studied. Some periplasmic proteins participate in nutrient acquisition. Others involved in energy conservation.

The outer membrane lies outside the thin peptidoglycan layer and is thought to be linked to the cell in two ways.
- First is by Braun's lipoprotein, the most abundant protein in the outer membrane. This small lipoprotein is covalently joined to the underlying peptidoglycan and is embedded in the outer membrane by its hydrophobic end.
- The second linking mechanism not as widely accepted. It involves contact sites that appear to join the outer membrane and the plasma membrane. These contact sites not observed when cells are fixed by rapid freezing, and it's argued that they may be artifacts of chemical fixation.

Most unusual constituents of the outer membrane are its lipopolysaccharides.

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Lipopolysaccharides (LPSs) Structure

Large, complex molecules that contain both lipid and carbohydrate, and consist of three parts. (1) Lipid A, (2) the core polysaccharide, and (3) the O side chain.

The lipid A region contains two glucosamine sugar derivatives, each with 3 fatty acids and phosphate or pyrophosphate attached. The fatty acids of lipid A are embedded in the outer membrane, while the remainder of the LPS molecule projects from the surface.

The core polysaccharide is joined to lipid A. The O side chain or O antigen is a polysaccharide chain extending outward from the core. It has several peculiar sugars and varies in composition between bacterial stains.

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Lipopolysaccharides (LPSs) Function

Many important functions.

1) Contributes to the negative charge on the bacterial surface because the core polysaccharide usually contains charged sugars and phosphate.

2) Helps stabilize outer membrane structure because lipid A is a major constituent of the exterior leaflet of the outer membrane.

3) May contribute to bacterial attachment to surfaces and biofilm formation.

4) Helps create a permeability barrier. The geometry of LPS and interactions between neighboring LPS molecules are thought to restrict the entry of bile salts, antibiotics, and other toxic substances that might kill or injure the bacterium.

5) Also plays a role in protecting pathogenic gram- bacteria from host defenses. The O side chain of LPS is also called the O antigen because it elicits an immune response by an infected host. This response involves the production of antibodies that bind the strain-specific form of LPS that elicited the response. However, many gram- bacteria can rapidly change the antigenic nature of their O side chains, thus thwarting host defenses.

6) Importantly, the lipid A portion of LPS is toxic; as a result, LPS can act as an endotoxin and cause some of the symptoms that arise in gram- bacterial infections. If LPS or lipid A enters the bloodstream, a form of septic shock develops, for which there's no direct treatment.

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Mechanism of Gram Staining

The difference between those bacteria that stain gram+ and those that stain gram- is due to the physical nature of their cell walls. If the cell wall is removed from gram+ bacteria, they stain gram-. Furthermore, bacteria that never make cell walls, such as the mycoplasmas, also stain gram-.

During the procedure, bacteria are first stained with crystal violet and next treated with iodine to promote dye retention. When bacteria are treated ethanol in the decolorization step, the alcohol is thought to shrink the pores of the thick peptidoglycan found in the cell walls of most gram+ bacteria, causing the peptidoglycan to act as a permeability barrier that prevents loss of crystal violet. Thus the dye-iodine complex is retained during the decolorization step and the bacteria remain purple.

In contrast, the peptidoglycan in gram- cell walls is very thin, not as highly crosslinked, and has larger pores. Alcohol treatment also may extract enough lipid from the outer membrane to increase the cell wall's porosity further. For these reasons, alcohol more readily removes the crystal violet-iodine complex from gram- bacteria. Thus gram- bacteria are easily stained red or pink by the counterstain safranin.

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Porin Proteins

Despite role of LPS in creating a permeability barrier, the outer membrane is more permeable than the plasma membrane and permits the passage of small molecules such as glucose and other monosaccharides. This is due to porin proteins.

Most of them cluster together to form a trimer in the outer membrane. Each porin protein spans the outer membrane and is more or less tube shaped; its narrow, water-filled channel allows passage of molecules smaller than about 600 daltons. However, large molecules such as vitamin B12 also cross the outer membrane. Such large molecules don't pass through porins, but have specific carries transport them instead.

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Osmotic Protection

Microbes have several mechanisms to changes in osmotic pressure. This pressure arises whent he concentration of solutes inside the cell differs from that outside, and the responses work to equalize the solute concentrations. However, in certain situations, osmotic pressure can exceed the cell's ability to acclimate. In these cases, additional protection is provided by the cell wall. When cells are in hypotonic solutions-ones in which the solute concentration is less than that in the cytoplasm-water diffuses into the cell, causing it to swell. Without the cell wall, the pressure on the plasma membrane would become so great that the membrane would be disrupted and the cell would burst-a process called lysis. Conversely, in hypertonic solutions, water flows out and the cytoplasm shrivels up-a process called plasmolysis.

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Cell Wall Protection

The protective nature of the cell wall is most clearly demonstrated when bacterial cells are treated with lysozyme or pencillin.

The enzyme lysozyme attacks peptidoglycan by hydrolyzing the glycosidic bond that connects N-acetylmuramic acid with N-acetylglucosamine.

Pencillin works by a different mechanism. It inhibits the enzyme transpeptidase, which is responsible for making the cross-links between peptidoglycan chains.

If bacteria are treated with either of these substances while in a hypotonic solution, they lyse. However, if in an isotonic solution, they survive and grow normally.

If they're gram+, treatment with lysozyme or penicillin results in the complete loss of the cell wall, and the cell becomes a protoplast.

When gram- bacteria are exposed to lysozyme or penicillin, the peptidoglycan layer is lost, but the outher membrane remains. These cells are called spheroplasts. Because they lack a complete cell wall, both protoplasts and spheroplasts are osomotically sensitive. If they are transferred to a hypotonic solution, they lyse.