PART 1 Introduction
2.1. ARE ALL CELLS THE SAME?
2.1.4. Procaryotes
The sizes of most procaryotes vary from 0.5 to 3 micrometers (mm) in equivalent radius.
Different species have different shapes, such as spherical or coccus (e.g., Staphylococci), cylindrical or bacillus (E. coli), or spiral or spirillum (Rhodospirillum). Procaryotic cells grow rapidly, with typical doubling times of one-half hour to several hours. Also, procary- otes can utilize a variety of nutrients as carbon source, including carbohydrates, hydrocar- bons, proteins, and CO2.
2.1.4.1. Eubacteria. The Eubacteria can be divided into several different groups. One distinction is based on the gram stain (developed by Hans Christian Gram in 1884). The staining procedure first requires fixing the cells by heating. The basic dye, crystal violet, is added; all bacteria will stain purple. Next, iodine is added, followed by
Figure 2.1. Replication of a virulent bacteriophage. A virulent phage undergoes a lytic cycle to produce new phage particles within a bacterial cell. Cell lysisreleases new phage particles that can infect more bacteria. (With permission, from J. G. Black, Microbiology:
Principles and Applications,3d ed., p. 282. This material is used by permission of John Wiley & Sons, Inc.)
the addition of ethanol. Gram-positivecells remain purple, while gram-negativecells be- come colorless. Finally, counterstaining with safranin leaves gram-positive cells purple, while gram-negative cells are red. This ability to react with the gram stain reveals intrinsic differences in the structure of the cell envelope.
A typical gram-negative cell is E. coli(see Fig. 2.2). It has an outer membranesup- ported by a thin peptidoglycan layer. Peptidoglycan is a complex polysaccharide with amino acids and forms a structure somewhat analogous to a chain-link fence. A second membrane (the inner or cytoplasmic membrane) exists and is separated from the outer membrane by the periplasmic space. The cytoplasmic membrane contains about 50% pro- tein, 30% lipids, and 20% carbohydrates. The cell envelope serves to retain important cellular compounds and to preferentially exclude undesirable compounds in the environ- ment. Loss of membrane integrity leads to cell lysis(cells breaking open) and cell death.
The cell envelope is crucial to the transport of selected material in and out of the cell.
A typical gram-positive cell is Bacillus subtilis. Gram-positive cells do not have an outer membrane. Rather they have a very thick, rigid cell wall with multiple layers of pep- tidoglycan. Gram-positive cells also contain teichoic acidscovalently bonded to the pepti- doglycan. Because gram-positive bacteria have only a cytoplasmic membrane, they are often much better suited to excretion of proteins. Such excretion can be technologically advantageous when the protein is a desired product.
Some bacteria are not gram-positive or gram-negative. For example, the Myco- plasmahave no cell walls. These bacteria are important not only clinically (e.g., primary atypical pneumonia), but also because they commonly contaminate media used industri- ally for animal cell culture.
16 An Overview of Biological Basics Chap. 2
Figure 2.2. Schematic of a typical gram-negative bacterium (E. coli). A gram-positive cell would be similar, except that it would have no outer membrane, its peptidoglycan layer would be thicker, and the chemical composition of the cell wall would differ signifi- cantly from the outer envelope of the gram-negative cell.
Actinomycetes are bacteria, but, morphologically, actinomycetes resemble molds with their long and highly branched hyphae. However, the lack of a nuclear membrane and the composition of the cell wall require classification as bacteria. Actinomycetes are impor- tant sources of antibiotics. Certain Actinomycetes possess amylolytic and cellulolytic en- zymes and are effective in enzymatic hydrolysis of starch and cellulose. Actinomyces, Thermomonospora, and Streptomycesare examples of genera belonging to this group.
Other distinctions within the eubacteria can be made based on cellular nutrition and energy metabolism. One important example is photosynthesis. The cyanobacteria (for- merly called blue-green algae) have chlorophyll and fix CO2 into sugars. Anoxygenic photosynthetic bacteria (the purple and green bacteria) have light-gathering pigments called bacteriochlorophyll. Unlike true photosynthesis, the purple and green bacteria do not obtain reducing power from the splitting of water and do not form oxygen.
When stained properly, the area occupied by the procaryotic cell’s DNA can be eas- ily seen. Procaryotes may also have other visible structures when viewed under the micro- scope, such as ribosomes, storage granules, spores, and volutins. Ribosomes are the site of protein synthesis. A typical bacterial cell contains approximately 10,000 ribosomes per cell, although this number can vary greatly with growth rate. The size of a typical ribo- some is 10 to 20 nm and consists of approximately 63% RNA and 37% protein. Storage granules (which are not present in every bacterium) can be used as a source of key metabolites and often contain polysaccharides, lipids, and sulfur granules. The sizes of storage granules vary between 0.5 and 1 mm.
Some bacteria make intracellular spores (often called endospores in bacteria). Bac- terial spores are produced as a resistance to adverse conditions such as high temperature, radiation, and toxic chemicals. The usual concentration is 1 spore per cell, with a spore size of about 1 mm. Spores can germinate under favorable growth conditions to yield ac- tively growing bacteria.
Volutin is another granular intracellular structure, made of inorganic polymetaphos- phates, that is present in some species. Some photosynthetic bacteria, such as Rhodospir- illum, have chromatophores that are large inclusion bodies (50 to 100 nm) utilized in photosynthesis for the absorption of light.
Extracellular products can adhere to or become incorporated within the surface of the cell. Certain bacteria have a coating or outside cell wall called capsule, which is usu- ally a polysaccharide or sometimes a polypeptide. Extracellular polymers are important to biofilm formation and response to environmental challenges (e.g., viruses). Table 2.3 summarizes the architecture of most bacteria.
2.1.4.2. Archaebacteria. The archaebacteria appear under the microscope to be nearly identical to many of the eubacteria. However, these cells differ greatly at the molecular level. In many ways the archaebacteria are as similar to the eucaryotes as they are to the eubacteria. Some examples of differences between archaebacteria and eubac- teria are as follows:
1. Archaebacteria have no peptidoglycan.
2. The nucleotide sequences in the ribosomal RNA are similar within the archaebacte- ria but distinctly different from eubacteria.
3. The lipid composition of the cytoplasmic membrane is very different for the two groups.
18 An Overview of Biological Basics Chap. 2 TABLE 2.3 Characteristics of Various Components of Bacteria
Part Size Composition and comments
Slime layer
Microcapsule 5–10 nm Protein–polysaccharide–lipid complex responsible for the specific antigens of enteric bacteria and other species.
Capsule 0.5–2.0 mm Mainly polysaccharides (e.g., Streptococcus); some- times polypeptides (e.g., Bacillus antracis).
Slime Indefinite Mainly polysaccharides (e.g., Leuconostoc); some- times polypeptides (e.g., Bacillus subtilis).
Cell wall
Gram-positive 10–20 nm Confers shape and rigidity upon the cell. 20% dry
species weight of the cell. Consists mainly of macromole-
cules of a mixed polymer of N-acetyl muramic- peptide, teichoic acids, and polysaccharides.
Gram-negative 10–20 nm Consists mostly of a protein–polysaccharide–lipid
species complex with a small amount of the muramic
polymer.
Cell membrane 5–10 nm Semipermeable barrier to nutrients. 5% to 10% dry weight of the cell, consisting of 50% protein, 28%
lipid, and 15% to 20% carbohydrate in a double- layered membrane.
Flagellum 10–20 nm by 4–12 mm Protein of the myosin–keratin–fibrinogen class, MW of 40,000. Arises from the cell membrane and is responsible for motility.
Pilus(fimbria) 5–10 nm by 0.5–2.0 mm Rigid protein projections from the cell. Especially long ones are formed by Escherichia coli.
Inclusions
Spore 1.0–1.5 mm by 1.6–2.0 mm One spore is formed per cell intracellularly. Spores show great resistance to heat, dryness, and antibacterial agents.
Storage granule 0.5–2.0 mm Glycogenlike, sulfur, or lipid granules may be found in some species.
Chromatophore 50–100 nm Organelles in photosynthetic species. Rhodospirillum rubrumcontains about 6000 per cell.
Ribosome 10–30 nm Organelles for synthesis of protein. About 10,000 ribosomes per cell. They contain 63% RNA and 37% protein.
Volutin 0.5–1.0 mm Inorganic polymetaphosphates that stain metachro- matically.
Nuclear material Composed of DNA that functions genetically as if the genes were arranged linearly on a single end- less chromosome, but that appears by light mi- croscopy as irregular patches with no nuclear membrane or distinguishable chromosomes. Au- toradiography confirms the linear arrangement of DNA and suggests a MW of at least 1000 ¥106. With permission, from S. Aiba, A. E. Humphrey, and N. F. Millis, Biochemical Engineering, 2d ed., Univer- sity of Tokyo Press, Tokyo, 1973.
The archaebacteria usually live in extreme environments and possess unusual me- tabolism. Methanogens, which are methane-producing bacteria, belong to this group, as well as the thermoacidophiles. The thermoacidophiles can grow at high temperatures and low pH values. The halobacteria, which can live only in very strong salt solutions, are members of this group. These organisms are important sources for catalytically active proteins (enzymes) with novel properties.