1. The eukaryotic and procaryotic cell structure.

2. Biological Domains: The domains Bacteria (cap. B and set in italics, also known as the eubacteria), Archaea, and Eukarya are based on 16S (bacteria) and 18S (eucaryotic) ribosomal RNA sequence data and comparison. What is the advantage of this method over the hierarchaical system for determining evolutionary relatedness? Why are the Archaea of so much interest to astrobiologists?

3. Naming of Bacteria: Uses the binomial system of nomenclature. Name given to an organism is derived from Latin or Greek and reflect a property(ies) of an organism, and occasionally an organism is named to honor someone. All organisms given two names;

• genus- first letter capitalized and word italicized or underlined.
• species- first letter lower cased and word underlined or italicized. Several species may make up a genus.
  • If two members of the same species differ slightly, but not enough to merit a new species designation, they are called variants or strains of a species. Are all E. coli pathogenetic?

4. Properties of microscopes that allow visualization of microorganisms:

• magnification power- how large an object is made to appear.
  • light microscope- 10X-2,000X
  • electron microscope- 500X- ~100,000X
• resolution- the distance that two objectives can be brought together and still be seen as two distinct objectives.
  • light microscope- 0.2 mm (with oil; 0.42 mm without oil)
  • electron microscope (EM)- 0.003 mm
    
• viruses can only be seen with the use of an electron microscope. Why?

5. Properties of bacteria as determined by light microscopy:

• enhancing contrast-
  • can use special types of microscopes or stains.
  • The Gram Stain, a differential stain (procedure- see fig 3.13 and 3.14)
    • Significance- allows separation of the Bacteria (eubacteria) into two groups, Gram-positive (purple staining) and Gram-negative (red-staining) bacteria. Gram staining is one of the first steps in the identification of an unknown bacterium. Also, the Gram reaction of eubacteria correlates with the structure/chemistry of cell wall. However, two bacteria that share the same Gram reaction may not be the same species!
      
• size- 0.2 mm to 40 mm, but larger bacteria have been reported.
  
• shape and size- genetically determined.
  • most common shapes- sphere, rod, comma, and helical.
  • some bacteria characterized by their absence of a consistent cell shape (pleomorphic (having many shapes)- bacteria of the genus Mycoplasma).
    
• cell groupings- following cell division, cells may remain together, depending on the species and growth conditions, to give rise to characteristic cell groupings- pairs, chains, grape-like clusters, or packets of 4 or 8 cells (Fig. 3.22). Why are terms like Straphylococcus, Streptococcus, or Bacillus not appropriate in referring to specific cell arrangements?
  
• motility- motility is most often associated with the presence of the structure known as the bacterial flagellum(a). Not all bacteria are motile. In some cases, cell movement is observed in the absence of a flagellar structure(s) (gas vesicles, see below). Can not see flagellum(a) with light microscope, only see cell(s) move.

6. Ultrastructure of a bacterial cell (structures can only be visualized with the electron microscope):

Bacterial cell can be separated into two compartments; the cell envelope (capsule and slime layer, cell wall layers, and cytoplasmic membrane) and its appendages (fimbriae/pili and flagellum(a)), and the cytosol (components within cytoplasm). Not all structures discussed will be present in all bacterial species.

The cell envelope is important in that (1) one of the structures, the cytoplasmic membrane is responsible for energy production that we generally associate with an intracellular organelle in eucaryotic systems, namely the mitochondria or the chloroplasts in plants, (2) protects the bacterial cell against chemical and biological threats that may be present in the environment, and (3) together with its appendages make possible the colonization, the attachment and growth, of bacteria to surfaces.

Cell envelope:

• Cytoplasmic membrane:
  
  • chemistry- composed of a phospholipid bilayer. Inserted into this layer are a large array of proteins.
    
  • two major functions-
    
    • energy transformation- In the case of organisms that grow by respiration or photosynthesis, chemical or radient energy is concerted into electrochemical energy or proton motive force. Proton motive force can be used to (1) turn bacterial flagellum, (2) transport certain molecule into cell, or (3) used to make chemical energy in form of ATP. In eukaryotes, proton motive force is established across inner membrane of mitochondria or chloroplasts.
      
    • transport- allows materials to be selectively transported across cytoplasmic membrane (membrane is a semi-primeable membrane). Mechanisms-
      
      • simple (no membrane protein involved) and facilitated diffusion (membrane protein involved) require no expenditure of energy. Concentraion of molecule being transported must be higher on outside than inside of cell. Concentration of material inside cell can never be higher than exists outside cell. Give an example of a molecule that enters by simple and facilitated diffusion. Fig. 3.27
        
      • active transport- membrane proteins involved, and process requires an expenditure of metabolic energy (proton motive force or chemical energy). Allows concentration of a molecule inside cell. How do ABC transport systems differ from the major facilitator transport systems? Fig. 3.28 and 3.29
        
      • Group translocation- What is the difference between active transport and group translocation? Fig. 3.30
        
      • one transport mechanism common to many eukaryotes but not bacteria is by the process known as "cytosis." Phagocytosis is an example.
        
• Bacteria lacking a cell wall structure: The mollicutes group (Mycoplasma). This is the only group wiith the domain Bacteria that lack a cell wall and contain sterols in their cytoplasmic membrane. Sterols, like cholesterol, are commonly found in animal membranes. Can one treat pneumonia caused by Mycoplasma pneumoniae with penicillin and if not why? See below
  
• cell wall (domain Bacteria, eubacteria):
  
  • Structure-
    • Gram-positive wall: appears as a single layered structure (EM) and is composed primarily of many layers of peptidoglycan.
      
    • Gram-negative wall: appears as a two layer structure. Inner layer composed of entirely of peptidoglycan (mono or bilayer), and an outer cell wall layer complex in its chemistry (phospholipid, lipoprotein, protein, and lipopolysaccharide). Region between this layer and cytoplasmic membrane is called the periplasmic space which contains a variety of enzymes, proteins, and small molecules essential for entry of certain molecules into the cell, and cell function.
      
  • chemistry- peptidoglycan, unique to Bacteria. Fig. 3.32
    
    • composed of two repeating sugars, N-acetylglucosamine (NAG), and N-acetylmuramic acid (NAM).
      
    • sugars held together by a covalent bond to form a peptidoglycan strand (many repeating sugar subunits).
      
    • adjacent peptidoglycan strands are held together by a short polypeptide chains that are covalently linked to NAM.
      
    • mode of action of the family of antibiotics known as the b-lactams (penicillin's) is to prevent crosslinking from occurring. For these antibiotics to be effective bacteria must be actively sythesizing peptidoglycan.
      
  • chemistry- outer cell wall layer (OCWL) of Gram-negative bacteria. Fig. 3.34
    
    • backbone structure- consist of a monolayer of phospholipid and a layer of lipopolysaccharide (LPS). LPS- 2 points I would like you to know are (1) the portion of LPS that allows association with the hydrophobic part of the lipid bilayer is called lipid A (hydrophobic), and it is this part of LPS that is responsible for some of the symptoms we associate with Gram-negative infections, shock, fever, etc., and (2) the part of the LPS molecule that extends from cell surface is called the O-specific side chain. This region is composed of a unique array of sugars, which can vary from one Gram-negative species to another and even within a species, and give a unique chemistry to LPS which we can used in the identification of Gram-negative bacteria. What is endotoxin?
    • porins (proteins which form holes or channels through OCWL)- allow passage of small molecules across this hydrophobic layer.
      
  • function- provides shape and rigidity (allows cells to live in dilute solute environments). If treat cells with lysozyme, and enzyme found in egg whites and many body fluids, peptidoglycan is digested, and cell loses shape and lysis. Lipopolysaccharide may also help in avoiding phagocytosis. What is the mechanism of action of lysozyme?
    
• Cell wall the Archaea:
  structure and chemistry is much more complex than that of the Bacteria (eubacteria), and in all cases no peptidoglycan is present.
  
• glycocalyx layer:
  • outer most layer of bacterial cell if present.
    
  • chemistry can vary from species to species, and even within a species. In most cases is carbohydrate in composition (composed of one or more sugars) or glycoprotein in composed (sugars covalently liked to protein), but in two cases the glycocalyx layer has a protein chemistry (B. anthracis and B. megaterium ) composed of a single amino acid, glutamic acid, and the D-isomer of the amino acid!. Most proteins made from 20 different amino acids, and all are of the L-isomer type. See Fig. 2.14
    
  • can identify bacteria based on chemistry of layer (chemical analysis and/or serotyping). What is the basic principal of serotyping?
    
  • role or function- Under most laboratory conditions layer is not essential to growth or viability of bacterium, but may alter properties (how colonies appear on surface of an agar medium). The two functions ascribed to the glycocalyx layer are protection (helps cells avoid phagocytosis, and desiccation), and attachment. In some cases, synthesis is dependent on environment (S. mutans makes structure only when grown on table sugar, sucrose). Glycocalyx layer may contribute to pathogenicity of bacterium.
    

Protein appendages:

• flagellum(a): ~1/2 of all bacteria are motile by means of a flagellum. Genetically determined.
  
  • arrangements on cell- montrichously, lophotrichously, and peritrichously flagellated.
    
  • structure- all flagella have a similar structure (Fig. 3.38). Three component parts are: filament (composed on identical protein subunits called flagellin whose chemistry varies from species to species, i.e., amino acid sequence of protein is different. Filament can be many times the length of the cell); hook (swivel on which filament turns); and basal body structure (anchors flagellum to cell by association with cytoplasmic membrane).
    
  • function- generates motion by having structure(s) rotate in either a clockwise or counterclockwise direction (if more than one flagellum on a cell all rotate in same direction). Process requires an expenditure of energy which is provided by proton motive force. The eukaryotic flagellum/cilia (structure different) moves up and down, and is a ATP dependent process.
    
• fimbriae/pili:
  
  • structure (Fig. 3.41)- appear as "hair-like" structures originating from cell surface (much thinner, short, and in some cases more numerous than flagella). A cell may have different types of fimbriae on same cell, and the number of copies of structures may vary.
    
  • chemistry- composed of identical protein subunits called pilin. Chemistry of pilin can vary from one type of pili to another, and can do serotyping using the pilin antigen.
    
  • function- attachment or adherence (may contribute to pathogenicity of a bacterium), and in case of sex-pilus structure required for conjugation (a process that allows transfer of DNA from one cell to another).

Cytosol

• nucleoid region:
  
  • most often consist of a single double stranded closed circular molecule of DNA. Some bacteria have been shown to have a linear double strand DNA genome like that of eukaryotes. Fig. 3.42
    
  • in general, have a single gene for every trait. Thus, bacteria are haploid organisms.
    
  • not enclosed in a membrane structure as we have in eukaryotic cells.
    
  • plasmids: small (about 1/10 size of chromosome) DNA molecules that contain additional genetic information not present on chromosome. A single bacterial cell can have several different plasmids (carrying different genetic information), and each plasmid type may vary in the number of copies per cell. A group of plasmids called R-plasmids (R-resistant plasmids) contain genes that encode for antibiotic resistance.
    
• Ribosome's (70S): Fig. 3.43
  
  • cell may contain from several hundred to several thousand ribosome's depending on phase of growth.
    
  • 70S ribosome composed of two subunits: a 50S and a 30S subunits. In contrast, the eukaryotic ribosome, 80S, is composed of a 60S and 40S subunit.
    
  • function- site of protein synthesis. The ribosome is the site of antibiotics like tetracycline, streptomycin, kanamycin, chloramphenicol, etc. Why do these antibiotics have no effect on eucaryotic protein synthesis?
    
• Endospores: Fig. 3.46
  • Property of only certain bacteria (genus Bacillus and Clostridium).
  • A servival mechanism
  • properties of endospore are different than vegetative, growing, cell.
  • endospores can differentiate back into a vegetative cell when conditions are suitable for growth.
• Eukaryotes
  • Endosymbiotic Theory: mitochondria and chloroplasts evolved from bacteria.
  • protein localization: transport of proteins to certain organelles.

Learning Objectives:

• What is the basis for recognizing two bacterial domains or kingdoms?
• What general features would allow you to distinguish between a procaryotic and a eucaryotic cell?
• How are bacteria named?
• Define resolution and magnification.
• Describe the Gram Stain, and what is the significance of this staining procedure.
• Identify the basic size and shapes of bacteria, and the cell arrangement possible following cell division.
• What is the glycocalyx layer, and what is its role?
• Compare the procaryotic and eucaryotic flagellum with regard to its structure, composition, and the way movement is generated.
• What are pili/fimbriae, and what is their role or purpose?
• Compare and contrast the cell walls of Gram-positive and Gram- negative bacteria. What unique feature distinguish the archaebacteria and the mycoplasmas with regard to the cell wall?
• How does the bacterial cytoplasmic membrane differ from the eucaryotic cell membrane with respect to chemical structure and function.
• Describe the various mechanisms that allow passage of substances across the cytoplasmic membrane.
• How do structures like the bacterial chromosome and ribosomes differ from their eucaryotic counterparts.
• What are plasmids, and how important are they to the bacterium?
• What types of reserve materials are accumulated/synthesized by bacteria?

 

6/1014/2009