Chapter 03 Lecture Outline PDF

Chapter 03 Lecture Outline See separate PowerPoint slides for all figures and tables pre- inserted into PowerPoint without notes. ©McGraw-Hill Education  A Glimpse of History Hans Christian Joachim Gram (1853 to 1...

Chapter 03 Lecture Outline See separate PowerPoint slides for all figures and tables pre- inserted into PowerPoint without notes. ©McGraw-Hill Education  A Glimpse of History Hans Christian Joachim Gram (1853 to 1938) Danish physician working at morgue in Berlin Worked for Dr. Carl Friedlander Attempting to identify cause of pneumonia Gram was developing methods to stain bacteria With one method, bacteria stained unequally Some retained dye, others did not Revealed two different kinds of bacteria Basis for modern Gram stain Identifies two major groups of bacteria according to cell wall structure and chemistry Gram-positive and Gram-negative ©McGraw-Hill Education  Microscopy and Cell Structure Microscopy reveals two fundamental cell types: Prokaryotic cells (Bacteria, Archaea) Smaller size gives high surface area to low volume Facilitates rapid uptake of nutrients, excretion of wastes Allows rapid growth Disadvantages include vulnerability to threats including predators, parasites, and competitors Eukaryotic cells (Eukarya) Larger, more complex, many cellular processes take place in membrane-bound compartments Defined by the presence of a nucleus ©McGraw-Hill Education  Microscopes Light microscope can magnify 1,000x Common, important tool in microbiology Electron microscope (1931) can magnify more than 100,000x Scanning probe microscope (1980s) can produce images of individual atoms on a surface ©McGraw-Hill Education  Principles of Light Microscopy (1) Light passes through specimen and then series of magnifying lenses Bright-field microscope is most common type Three key concepts: Magnification: apparent increase in size Resolution: resolving power, or ability to distinguish two objects that are very close together Contrast: difference in color intensity between an object and the background; determines how easily cells can be seen ©McGraw-Hill Education  Principles of Light Microscopy (2) Magnification: apparent increase in size Modern compound microscope has two lens types: objective and ocular Magnification is product of objective (4x, 10x, 40x, or 100x) and ocular lens (10x) Condenser lens (between light source and specimen) focuses light on specimen; does not magnify ©McGraw-Hill Education  Principles of Light Microscopy – Figure 3.1 ©McGraw-Hill Education © Scenics & Science/Alamy Stock Photo  Principles of Light Microscopy (3) Resolution: resolving power, or ability to distinguish two objects that are very close together Defined as minimum distance between two points at which those points can be observed as separate Depends on quality and type of lens, wavelength of light, magnification, and specimen preparation Maximum resolving power of light microscope is 0.2 micrometer ©McGraw-Hill Education  Principles of Light Microscopy – Figure 3.2 Immersion oil used to displace air between lens and specimen when using high powered 100x objective Has same refractive index (measure of speed of light passing through medium) as glass Prevents refraction of light; keeps rays from missing opening in objective lens ©McGraw-Hill Education © McGraw-Hill Education/Lisa Burgess, photographer  Principles of Light Microscopy – Figure 3.3 Contrast determines how easily cells can be seen Transparent bacteria lack contrast, difficult to see against colorless background Stains increase contrast but kill microbes ©McGraw-Hill Education © McGraw-Hill Education/Lisa Burgess, photographer Light Microscopes That Increase Contrast – Figure 3.4 Dark-Field Microscope Cells appear as bright objects against dark background Directs light toward specimen at angle Only light scattered by specimen enters objective lens ©McGraw-Hill Education © McGraw-Hill Education/Lisa Burgess, photographer Light Microscopes That Increase Contrast – Figure 3.5 Phase-Contrast Microscope Special optics amplify difference between refractive index of dense material and surrounding medium Makes cells and other dense material appear darker ©McGraw-Hill Education © McGraw-Hill Education/Lisa Burgess, photographer  Light Microscopes That Increase Contrast – Figure 3.6 Differential Interference Contrast (DIC) Microscope Like phase-contrast, has special optics that depend upon differences in refractive index Separates light into two beams that pass through specimen and recombine Light waves are out of phase when recombined, yield three- dimensional appearance of image ©McGraw-Hill Education © Gerd Guenther/Science Source  Light Microscopes That Detect Fluorescence – Figure 3.7 Fluorescence Microscope Used to observe cells or materials either naturally fluorescent or tagged with fluorescent dyes Molecules absorb light at one wavelength (usually ultraviolet light) and emit light at longer wavelength Most today are epifluorescent: UV light projected onto, not through, specimen ©McGraw-Hill Education © Evan Roberts  Light Microscopes That Detect Fluorescence – Figure 3.8 Scanning Laser Microscope (SLM) Specimens stained with fluorescent dye Fluorescent molecules bind to certain internal compounds; marks their precise location Allows detailed interior views of intact cells May provide 3-dimensional images of thick structures ©McGraw-Hill Education Courtesy of A. Harrer, B. Pitts, P. Stewart/MSU-CBE  Light Microscopes That Detect Fluorescence (1) Confocal Microscopy Laser beam illuminates a point on one vertical plane of specimen Mirrors scan laser beam across specimen, illuminating successive planes Each plane represents one fine slice of specimen Computer constructs 3-dimensional image Like a miniature computerized axial tomography (CAT) scan for cells ©McGraw-Hill Education  Light Microscopes That Detect Fluorescence (2) Two-photon Microscopy (multiphoton microscopy) Similar to confocal, but lower energy used Less damaging to cells; allows time-lapse images Light penetrates deeper to give interior views of relatively thick structures ©McGraw-Hill Education  Light Microscopes That Detect Fluorescence – Figure 3.9 Super-Resolution Microscope Can improve resolution down to 10 nanometers ©McGraw-Hill Education both: © Dr. Henrik Strah/Newcastle University  Electron Microscopes – Figure 3.10 Can clearly magnify objects Light Microscope Transmission Electron Microscope 100,000x Uses electromagnetic lenses, electrons, and a fluorescent screen to replace glass lenses, visible light, and the eye Image photographed as black- and-white electron photomicrograph May be artificially colored ©McGraw-Hill Education  Electron Microscopes (1) Light Microscope Transmission Electron Wavelength of electrons Microscope approximately 1,000 shorter than light Resolving power approximately 1,000-fold greater: approximately 0.3 nanometer ©McGraw-Hill Education  Electron Microscopes (2) Lenses and specimen must be in vacuum Air molecules would interfere with electrons Impossible to observe living specimens Results in large, expensive unit and complex specimen preparation Two major types: Transmission Electron Microscope (TEM) Scanning Electron Microscope (SEM) ©McGraw-Hill Education  Electron Microscopes – Figure 3.11 Transmission Electron Microscope (TEM): beam of electrons passes through specimen or scatters Depends on density of region: dark areas are dense Thin-sectioning used to view internal details, but process can distort cells Freeze-fracturing, freeze-etching reveal shape of internal structures Newer methods that reduce damage to cells include cryo-electron microscopy (cryo-EM), and cryo-electron tomography for 3-D images ©McGraw-Hill Education a: © Lee D. Simon/Science Source; b: © Dr. Tony Brain/Science Source  Electron Microscopes – Figure 3.12 Scanning Electron Microscope (SEM): beam of electrons scans over surface of specimen Used to observe surface details Surface coated with thin film of metal Electrons released from specimen are observed Yields 3-D effect ©McGraw-Hill Education © Dennis Kunkel Microscopy/SPL/Science Source  Scanning Probe Microscopes – Figure 3.13 Detailed images of surfaces Example: atomic force microscope (AFM) Resolving power much greater than that of EM Avoids special preparation required for EM Sharp probe moves across sample’s surface “Feels” bumps, valleys of atoms Laser measures motion, computer produces surface map ©McGraw-Hill Education Source: Dr. Mary Ng Lee, National University of Singapore/CDC  Summary of Microscopic Instruments – Table 3.1 (1) Table 3.1 A Summary of Microscopic Instruments and Their Characteristics Instrument Mechanism Comment Visible light passes through a series of Relatively easy to use; considerably less Light Microscopes lenses to produce a magnified image. expensive than electron and scanning probe microscopes. Bright-field Illuminates the field of view evenly and Most common type of microscope. generates a bright background. Dark-field Light is directed toward the specimen at an Makes unstained cells easier to see; angle. organisms stand out as bright objects against a dark background. Phase-contrast Increases contrast by amplifying differences Dense material appears darker than in refractive index. normal. ©McGraw-Hill Education All: © McGraw-Hill Education/Lisa Burgess, photographer  Summary of Microscopic Instruments – Table 3.1 (2) Instrument Mechanism Comment Light Microscopes Visible light passes through a series of Relatively easy to use; considerably less lenses to produce a magnified image. expensive than electron and scanning probe microscopes. Differential Two light beams pass through the The image of the specimen appears Interference specimen and then recombine. three-dimensional. contrast Fluorescence Projects ultraviolet light, causing Used to observe cells stained or tagged fluorescent molecules in the specimen to with a fluorescent dye. emit longer wavelength light. Scanning laser Mirrors scan a laser beam across Used to obtain a three-dimensional successive regions and planes of a image of a structure that has been specimen. From that information, a stained with a fluorescent dye; provides computer constructs an image. detailed sectional views of intact cells. Super-resolution Complex illumination mechanisms and Higher resolving power than that of a merged data are used to construct the conventional fluorescence microscope. image. ©McGraw-Hill Education From top to bottom: © Gerd Guenther/Science Source; © Evan Roberts; © A. Harrer, B. Pitts, P. Stewart/MSU-CBE; © Dr. Henrik Strah/Newcastle University  Summary of Microscopic Instruments – Table 3.1 (3) Instrument Mechanism Comment Electron Microscopes Electron beams are used in place of Can clearly magnify images 1000,000x. visible light to produce the magnified image. Transmission Transmits a beam of electrons through Complicated specimen preparation is a specimen. required. Scanning A beam of electrons scans back and Used for observing surface details; forth over the surface of a specimen. produces a three-dimensional effect. Scanning Probe Microscopes A physical probe is used to produce Produces a map showing the bumps detailed images of surfaces. and valleys of the sample’s surface. Atomic Force Probe moves in response to even the No special sample preparation slightest force between it and the required; produces a three- sample. dimensional effect. ©McGraw-Hill Education Top: © Lee D. Simon/Science Source; Middle: © Dennis Kunkel Microscopy/SPL/Science Source; Bottom: Source: Dr. Mary Ng Mah Lee, National University of Singapore/CDC  Preparing Specimens for Light Microscopy – Figure 3.14 Wet mount uses a drop of liquid specimen overlaid with a coverslip Allows observation of living organisms Can be difficult to see when they are colorless Smear involves drying and fixing specimen before staining to visualize ©McGraw-Hill Education  Preparing Specimens for Light Microscopy – Table 3.2 Table 3.2 A Summary of Stains and Their Characteristics Stain Characteristic Simple Stains A basic dye is used to stain cells. Easy way to increase the contrast between otherwise colorless cells and a colorless background. Differential Stains A multistep procedure is used to stain cells and distinguish one group of microorganisms from another. Gram Stain Used to separate bacteria into two major groups: Gram-positive and Gram-negative. The staining characteristics of these groups reflect a fundamental difference in the chemical structure of their cell walls. This is by far the most widely used staining procedure. Acid-fast stain Used to detect organisms that do not easily take up stains, particularly members of the genus Mycobacterium. Special Stains A special procedure is used to stain specific cell structures. Capsule stain The common procedure darkens the background, so the capsule stands out as a clear area surrounding the cell. Endospore stain Stains endospores, a type of dormant cell that does not readily take up stains. Endospores are produced by Bacillus and Clostridium species. Flagella stain The staining agent adheres to and coats the otherwise thin flagella, making them visible with the light microscope Fluorescent Dyes and Fluorescent dyes and tags absorb ultraviolet light and then emit light of a longer wavelength. Tags Fluorescent dyes Some fluorescent dyes bind to compounds found in all cells; others bind to compounds specific to only certain types of cells. Fluorescent tags Antibodies to which a fluorescent molecule has been attached are used to tag specific molecules. ©McGraw-Hill Education  Simple Staining Simple staining uses a single dye to stain the specimen Basic dyes carry positive charge Attracted to negatively charged cellular components Examples are methylene blue and crystal violet Acidic dyes can be used for negative staining Cells repel the negatively charged dye; colorless cells stand out against background Can be done as wet mount ©McGraw-Hill Education  Differential Staining Differential staining used to distinguish different groups of bacteria Gram stain most widely used for bacteria Two groups: Gram-positive bacteria and Gram-negative bacteria Reflects fundamental difference in cell wall structure ©McGraw-Hill Education  Gram Stain – Figure 3.15a Flood smear with primary stain Rinse and flood with iodine, a mordant that stabilizes the dye in the cell Rinse and briefly add alcohol, a decolorizing agent, to remove dye complex from Gram-negative cells Rinse and flood smear with counterstain that adds a different color to Gram-negative cells Jump to Gram Stain – Figure 3.15a Long Description ©McGraw-Hill Education  Differential Staining – Figure 3.15 Success of Gram stain relies upon length of time of decolorizing step and age of culture ©McGraw-Hill Education b: © McGraw-Hill Education/Lisa Burgess, photographer  Acid-Fast Stain – Figure 3.16 Acid-fast stain used to detect organisms that do not readily take up dyes Detects Mycobacterium species such as causative agents of tuberculosis and Hansen’s disease (leprosy) Cell wall contains high concentrations of mycolic acids Multi-step procedure Primary stain is concentrated red dye Acid-fast cells retain red dye after being flooded with acid-alcohol Methylene blue used as counterstain ©McGraw-Hill Education Source: Dr. George P. Kubica/CDC  Special Stains to Observe Cell Structures – Figure 3.17 Capsule stain allows observation of gel-like layer that surrounds some microbes Capsule stains poorly, so background is stained to make capsule visible India ink added to wet mount is common method ©McGraw-Hill Education Source: Dr. Leanor Haley/CDC  Special Stains to Observe Cell Structures – Figure 3.18 Endospore stain allows visualization of endospores, resistant dormant cells often formed by Bacillus and Clostridium Endospore resists Gram stain; appears as clear object Endospore stain uses heat to facilitate uptake of the primary dye malachite green by endospore Counterstain (usually safranin) colors other cells pink ©McGraw-Hill Education © McGraw-Hill Education/Lisa Burgess, photographer  Special Stains to Observe Cell Structures – Figure 3.19 Flagella stain uses a substance that makes the dye adhere to thin flagella, making them visible Presence and distribution of flagella can help in identification Peritrichous: flagella surround cell Polar: flagellum on one end ©McGraw-Hill Education both: Source: Dr. William A. Clark/CDC  Special Stains to Observe Cell Structures – Figure 3.20 Fluorescent Dyes and Tags Some dyes bind to structures in all cells Some are changed by cellular processes: can distinguish between living and dead cells Immunofluorescence uses fluorescent dye-antibody labels to tag a unique microbe protein ©McGraw-Hill Education a: Source: CDC; b: © Evan Roberts  Morphology of Prokaryotic Cells: Shapes – Figure 3.21a to b Two types most common: Coccus: spherical Rod: cylindrical Also called a bacillus Short rods sometimes called coccobacillus Coccus Rod (bacillus) ©McGraw-Hill Education a: ©SciMAT/Science Source; b: ©Dennis Kunkel Microscopy/SPL/ Science Source  Morphology of Prokaryotic Cells: Shapes – Figures 3.21c-e and 3.22 Variety of other shapes: Vibrio, spirillum, spirochete Pleomorphic (many shapes) Great diversity often found in low nutrient environments ©McGraw-Hill Education Far left: © James Staley; c: ©Dennis Kunkel Microscopy/SPL/Science Source; d: ©Dennis Kunkel Microscopy/SPL/ Science Source; e: ©Dennis Kunkel Microscopy/SPL/Science Source  Morphology of Prokaryotic Cells: Arrangements – Figure 3.23 Most prokaryotes divide by binary fission Cells often stick together following division to form characteristic groupings Examples: Neisseria gonorrhoeae (diplococci) Streptococcus (chains) Sarcina (cubical packets) Staphylococcus (grape-like clusters) ©McGraw-Hill Education a (top): ©Dennis Kunkel Microscopy/SPL/Science Source; a (bottom): ©BSIP SA/Alamy Stock Photo; b: Source: Betsy Crane/CDC; c: ©Eye of Science/Science Source  Multicellular Associations Some prokaryotes live as multicellular associations Myxobacteria form swarms of cells that glide over moist surfaces as a pack Collectively release enzymes and degrade organic material, including other bacterial cells When water or nutrients become limiting, cells form fruiting body visible to naked eye Most bacteria on surfaces in natural habitat form polymer-encased communities called biofilms ©McGraw-Hill Education  Prokaryotic Cells Surface layers are cell envelope Cytoplasmic membrane Cell wall Capsule (if present) Cytoplasm Nucleoid Location of chromosome Locomotor appendages (if present) ©McGraw-Hill Education  Prokaryotic Cells – Figure 3.24 Jump to Prokaryotic Cells – Figure 3.24 Long Description ©McGraw-Hill Education b: © Science Source  Prokaryotic Cells – Table 3.3 (1) Table 3.3 Typical Prokaryotic Cell Structures Structure Characteristics Structures Outside the Cell Wall Filamentous appendages Composed of protein subunits that form a helical chain. Flagella Provide the most common mechanism of motility. Pili Different types of pili have different functions. The common types, often called fimbriae, allow cells to adhere to surfaces. A few types are used for twitching or gliding motility. Sex pili are involved in DNA transfer. Capsules and slime layers Layers outside the cell wall, usually made of polysaccharide. Capsule Distinct and gelatinous. Allows bacteria to adhere to specific surfaces; allows some organisms to avoid the body’s defense systems and thus cause disease. Slime layer Diffuse and irregular. Allows bacteria to adhere to specific surfaces. Cell Wall Peptidoglycan provides rigidity to bacterial cell walls, preventing the cells from lysing. Gram-positive Thick layer of peptidoglycan that contains teichoic acids and lipoteichoic acids. Gram-negative Thin layer of peptidoglycan surrounded by an outer membrane. The outer layer of the outer membrane is lipopolysaccharide. ©McGraw-Hill Education  Prokaryotic Cells – Table 3.3 (2) Table 3.3 Typical Prokaryotic Cell Structures Structure Characteristics Cytoplasmic Membrane Phospholipid bilayer embedded with proteins. Surrounds the cytoplasm, separating it from the external environment. Also transmits information about the external environment to the inside of the cell. Internal Components DNA Carries the genetic information of the cell. Chromosome Carries the genetic information required by a cell. Typically a single, circular, double-stranded DNA molecule. Plasmid Generally carries only genetic information that may be advantageous to a cell in certain situations. Endospore A type of dormant cell that is extraordinarily resistant to heat, desiccation, ultraviolet light, and toxic chemicals. Cytoskeleton Protein framework involved in cell division and control of cell shape. Gas vesicles Small, rigid structures that provide buoyancy to a cell. Granules Accumulations of high-molecular-weight polymers, synthesized from a nutrient available in relative excess. Ribosomes Involved in protein synthesis. Two subunits, 30S and 50S, join to form the 70S ribosome. ©McGraw-Hill Education  The Cytoplasmic Membrane – Figure 3.25 Cytoplasmic membrane defines boundary of cell Phospholipid bilayer embedded with proteins Hydrophobic tails face in; hydrophilic tails face out Proteins serve numerous functions Selective gates Sensors of environmental conditions Enzymes Fluid mosaic model: proteins drift about in lipid bilayer ©McGraw-Hill Education  The Cytoplasmic Membrane Bacteria and Archaea have same general structure of cytoplasmic membranes Distinctly different phospholipid compositions Lipid tails of Archaea are not fatty acids Connected differently to glycerol ©McGraw-Hill Education  Permeability of Cytoplasmic Membrane – Figure 3.26 Cytoplasmic membrane is selectively permeable O2, CO2, N2, small hydrophobic molecules, and water pass freely Some cells facilitate water passage with aquaporins Other molecules must be moved across membrane via transport systems Pass through easily: Passes through: Do not pass through: Gases (O2, CO2, N2) Water Sugars Small hydrophobic lons molecules Amino acids ATP Macromolecules a) The cytoplasmic membrane is selectively permeable. Gases, small b) Aquaporins allow water to pass through the cytoplasmic membrane hydrophobic molecules, and water are the only substances that more easily. pass freely through the phospholipid bilayer. ©McGraw-Hill Education  Permeability of Cytoplasmic Membrane Simple Diffusion Movement from high to low concentration until equilibrium is reached Speed of diffusion depends on concentration The greater the difference in concentration on either side of a membrane, the higher the rate of diffusion ©McGraw-Hill Education  Permeability of Cytoplasmic Membrane – Figure 3.27 (1) Osmosis: Diffusion of water across a selectively permeable membrane due to unequal solute concentrations Water diffuses from high water concentration (low solute concentration) to low water concentration (high solute concentration) Water flows from hypotonic to hypertonic solution Water flows across a membrane toward the hypertonic solution. No net water flow between Hypotonic solution Hypertonic solution isotonic solutions ©McGraw-Hill Education  Permeability of Cytoplasmic Membrane – Figure 3.27 (2) Environment of prokaryotes are typically dilute (hypotonic) Water flows into the cell where cytoplasm is a concentrated solution (hypertonic) Cell wall prevents cell from bursting Jump to Permeability of Cytoplasmic Membrane – Figure 3.27 (2) Long Description ©McGraw-Hill Education  Cytoplasmic Membrane in Energy Transformation – Figure 3.28 Electron Transport Chain (ETC) embedded in cytoplasmic membrane Uses energy from electrons to move protons out of cell Creates electrochemical gradient across membrane Energy called proton motive force Harvested to drive ATP synthesis and some forms of transport, motility ©McGraw-Hill Education  Transport of Small Molecules Across Cytoplasmic Membrane – Figure 3.29 Most molecules must pass through proteins functioning as selective gates Transport systems: permeases or carriers Membrane-spanning Highly specific: carriers transport certain molecule type 1. A given transport protein 2. Binding of that molecule changes 3. The molecule is released on the recognizes a specific molecule. the shape of the transport protein. other side of the membrane. ©McGraw-Hill Education  Transport of Small Molecules Across Cytoplasmic Membrane - Figure 3.30 (1) Facilitated diffusion is a form of passive transport Movement down gradient; no energy required Not typically useful in low-nutrient environments a) Facilitated diffusion b) Active transport, using Active transport, using ATP as an c) Group translocation proton motive force as an energy source. A binding protein Transporter allows a substance to energy source. gathers the transported molecules. Transporter chemically move across the membrane, but alters the substance as only down its concentration Transporter uses energy (ATP or proton motive force) to move a it is transported across gradient. substance across the membrane against a concentration gradient. the membrane. ©McGraw-Hill Education  Transport of Small Molecules Across Cytoplasmic Membrane - Figure 3.30 (2) Active transport requires energy Moves material against concentration gradient Sometimes driven by proton motive force Example is efflux pump Sometimes driven by ATP (ABC transporter) a) Facilitated diffusion b) Active transport, using Active transport, using ATP as an c) Group translocation proton motive force as energy source. A binding protein Transporter allows a substance gathers the transported molecules. Transporter chemically an energy source. to move across the membrane, alters the substance as but only down its concentration Transporter uses energy (ATP or proton motive force) to move a it is transported across gradient. substance across the membrane against a concentration gradient. the membrane. ©McGraw-Hill Education  Transport of Small Molecules Across Cytoplasmic Membrane - Figure 3.30 (3) Group Translocation is common in bacteria Chemically alters compound during passage through cytoplasmic membrane Phosphorylation common Often used by bacteria to bring glucose into the cell a) Facilitated diffusion b) Active transport, using Active transport, using ATP as an c) Group translocation proton motive force as an energy source. A binding protein Transporter allows a substance to Transporter chemically energy source. gathers the transported molecules. move across the membrane, but alters the substance as it only down its concentration Transporter uses energy (ATP or proton motive force) to move a substance is transported across the gradient. across the membrane against a concentration gradient. membrane. ©McGraw-Hill Education  Transport of Small Molecules Across Cytoplasmic Membrane – Table 3.4 Table 3.4 Transport Mechanisms Used by Prokaryotic Cells Transport Mechanism Characteristics Facilitated Diffusion Rarely used by prokaryotes. Exploits a concentration gradient to move molecules; can only eliminate a gradient, not create one. No energy is used. Active Transport Energy is used to accumulate molecules against a concentration gradient. Transporters that use As a proton is allowed into the cell, another proton motive force substance is either brought along or expelled. ABC transporters ATP is used as an energy source. Binding proteins deliver a molecule to the transporter. Group Translocation The transported molecule is chemically altered as it passes into the cell. ©McGraw-Hill Education  Transport of Small Molecules Across Cytoplasmic Membrane – Figure 3.31 Protein secretion: active movement out of cell Examples: extracellular enzymes, external structures Polypeptides tagged for secretion via signal sequence of amino acids Jump to Transport of Small Molecules Across Cytoplasmic Membrane – Figure 3.31 Long Description ©McGraw-Hill Education  The Cell Wall of Prokaryotic Cells Cell wall is strong, somewhat rigid structure that prevents cell from bursting Distinguishes two main types of bacteria: Gram-positive Gram-negative ©McGraw-Hill Education  The Cell Wall of Prokaryotic Cells – Table 3.5 Table 3.5 Comparison of Features of Gram-Positive and Gram-Negative Bacteria Gram-Positive Gram-Negative Color of Gram-Stained Cell Purple Pink Representative Genera Bacillus, Staphylococcus, Streptococcus Escherichia, Neisseria, Pseudomonas Distinguishing Structures/Components Peptidoglycan Thick layer Thin layer Teichoic acids Present Absent Outer membrane Absent Present Lipopolysaccharide (endotoxin) Absent Present Porin proteins Absent (unnecessary because there is no Present; allow molecules to pass outer membrane) through outer membrane General Characteristics Sensitivity to penicillin Generally more susceptible (with notable Generally less susceptible (with exceptions) notable exceptions) Sensitivity to lysozyme Yes No ©McGraw-Hill Education Top left: © McGraw-Hill Education/Lisa Burgess, photographer  Peptidoglycan – Figure 3.32 Peptidoglycan found only in Chemical structure of N-acetylglucosamine (NAG) bacteria and N-acetylmuramic acid (NAM); the ring structure of each molecule is glucose. Alternating series of subunits form glycan chains N-acetylmuramic acid (NAM) N-acetylglucosamine (NAG) Glycan chains are composed of alternating subunits of NAG and NAM. They are cross-linked via Tetrapeptide chain links glycan their tetrapeptide chains to create peptidoglycan. chains Peptide interbridge in Gram- positive cells Interconnected glycan chains form a large sheet. Multiple connected layers create a three-dimensional molecule. ©McGraw-Hill Education  The Gram-Positive Cell Wall – Figure 3.33 Relatively thick peptidoglycan layer Teichoic acids extend above peptidoglycan layer Gel-like material below peptidoglycan layer ©McGraw-Hill Education c: © Egbert Hoiczyk  The Gram-Negative Cell Wall – Figure 3.34 Thin peptidoglycan layer Outside is unique outer membrane ©McGraw-Hill Education © Egbert Hoiczyk  The Gram-Negative Cell Wall (1) Outer membrane: outside layer is lipopolysaccharide (LPS) Signals immune system of invasion by Gram-negative bacteria Small levels elicit response to eliminate invader LPS is also called endotoxin Large amounts accumulating in bloodstream can be deadly Includes Lipid A Recognized by immune system Includes O antigen Can be used to identify species or strains ©McGraw-Hill Education  The Gram-Negative Cell Wall (2) Outer membrane blocks passage of many molecules including certain antimicrobial medications Small molecules and ions can cross via porins Secretion systems important in pathogenesis Between cytoplasmic membrane and outer membrane is periplasmic space Filled with gel-like periplasm Exported proteins accumulate unless specifically moved across outer membrane Binding proteins of ABC transport systems ©McGraw-Hill Education  Antibacterial Substances That Target Peptidoglycan Interference with peptidoglycan can weaken cell wall and allow cell to burst Penicillin interferes with peptidoglycan synthesis Prevents cross-linking of adjacent glycan chains Usually more effective against Gram-positive bacteria than Gram- negative bacteria Outer membrane of Gram-negative cells blocks access Derivatives have been developed that can cross Lysozyme breaks bonds linking glycan chains Enzyme found in tears, saliva, other body fluids Destroys structural integrity of peptidoglycan molecule Usually more effective against Gram-positive bacteria ©McGraw-Hill Education  Cell Wall Type and the Gram Stain Crystal violet stains inside of cell Gram-positive cell wall prevents crystal violet–iodine complex from being washed out Decolorizing agent dehydrates thick layer of peptidoglycan; desiccated state acts as barrier Solvent action of decolorizing agent damages outer membrane of Gram-negative cell wall Thin layer of peptidoglycan cannot retain dye complex ©McGraw-Hill Education  Cell Wall Type and the Gram Stain – Figure 3.35 Gram-positive cell wall Large crystal violet-iodine complexes form The decolorizing agent dehydrates the within the cytoplasm as a result of the thick layer of peptidoglycan, causing it to first two steps of the Gram stain. become a tight mesh-like structure that prevents the crystal violet-iodine complexes from being washed out of the cell. Gram-negative cell wall Large crystal violet-iodine complexes form The decolorizing agent damages the within the cytoplasm as a result of the outer membrane. The thin peptidoglycan first two steps of the Gram stain. layer cannot prevent the crystal violet- iodine complexes from being washed out of the cell. ©McGraw-Hill Education  Bacteria That Lack a Cell Wall – Figure 3.36 Some bacteria lack a cell wall Mycoplasma species are flexible Unaffected by penicillin and lysozyme Cytoplasmic membrane contains sterols that increase its strength ©McGraw-Hill Education © Don W. Fawcett/Science Source  Cell Walls of Archaea Members of Archaea have variety of cell walls Probably due to wide range of environments Includes extreme environments Archaea less well studied than Bacteria No peptidoglycan, but some have similar molecule pseudopeptidoglycan Many have S-layers that self-assemble Built from sheets of flat protein or glycoprotein subunits ©McGraw-Hill Education  Capsules and Slime Layers – Figure 3.37 Gel-like layer outside cell wall protects or allows attachment Capsule: distinct, gelatinous Slime layer: diffuse, irregular Most composed of glycocalyx (sugar shell), but some are polypeptides Once attached to a surface, cells can grow as biofilm Polymer-encased community Example: dental plaque Some capsules allow bacteria to evade host immune system ©McGraw-Hill Education a: © Scimat/Science Source; b: © Scimat/Science Source  Flagella – Figure 3.38 Flagella involved in motility Spin like propellers to move cell Some important in disease Helicobacter pylori Numbers and arrangements help with characterization of bacteria ©McGraw-Hill Education a: ©CAMR/A. Barry Dowsett/Science Source; b: ©Dennis Kunkel Microscopy/SPL/ Science Source  Flagella – Figure 3.39 Three parts of bacterial flagellum: Basal body: anchors to cell wall and cytoplasmic membrane Hook Filament: made up of flagellin subunits Archael flagella: Chemically distinct Use energy from ATP instead of proton motive force ©McGraw-Hill Education  Chemotaxis – Figure 3.40 Chemotaxis: Bacteria sense a chemical and move toward it (nutrient) or away from it (toxin) Movement is series of runs (straight line) and tumbles (changes in direction) due to coordinated rotation of flagella Jump to Chemotaxis – Figure 3.40 Long Description ©McGraw-Hill Education  Chemotaxis – Figure 3.41 Other responses observed: Aerotaxis Magnetotaxis Thermotaxis Phototaxis ©McGraw-Hill Education ©Dennis Kunkel Microscopy/SPL/Science Source  Pili – Figure 3.42 Pili (singular: pilus) are shorter than flagella Types that allow surface attachment also called fimbriae Twitching motility and gliding motility involve pili Sex pilus used to join bacteria for a type of DNA transfer ©McGraw-Hill Education a: ©Dennis Kunkel/SPL/Science Source; b: Source: Harley W. Moon/U.S. Department of Agriculture  Internal Components of Prokaryotic Cells – Figure 3.43 Chromosome forms gel-like region: the nucleoid Single circular double-stranded DNA molecule Packed tightly via binding proteins and supercoiling Plasmids Do not encode essential genetic information Similar structure to chromosome, but much smaller May be shared with other bacteria; antibiotic resistance can spread this way ©McGraw-Hill Education ©CNRI/SPL/Science Source  Internal Components of Prokaryotic Cells – Figure 3.44 Ribosomes involved in protein synthesis Facilitate joining of amino acids Relative size and density expressed as S (Svedberg unit) that reflects how fast they settle when centrifuged Prokaryotic ribosomes are 70S Composed of 30S and 50S subunits Eukaryotic ribosomes are 80S Important medically: antibiotics impacting 70S ribosome do not affect 80S ribosome ©McGraw-Hill Education  Internal Components of Prokaryotic Cells Cytoskeleton: interior protein framework Bacterial proteins similar to eukaryotic cytoskeleton have been characterized Likely involved in cell division and controlling cell shape Storage granules: accumulations of polymers Synthesized from nutrients available in excess Carbon, energy storage: ‐‐ Glycogen Poly β hydroxybutyrate  PHB  Metachromatic granules stain red with methylene blue Gas vesicles: controlled to provide buoyancy ©McGraw-Hill Education  Endospores – Figure 3.45 Endospores: unique type of dormant cell Produced by members of Bacillus, Clostridium May remain dormant for 100 years or longer Extremely resistant to heat, desiccation, chemicals, ultraviolet light, boiling water Endospores can germinate to become vegetative cells that can multiply Found virtually everywhere ©McGraw-Hill Education © Dr. Kari Lounatmaa/Science Source  Endospores Sporulation triggered by limited carbon or nitrogen Starvation begins 8-hour process Endospore layers prevent damage Exclude molecules (for example, lysozyme) Cortex maintains core in dehydrated state, protects from heat Core has small proteins that bind to and protect DNA Calcium dipicolinate seems to play important protective role Germination triggered by heat, chemical exposure Not a means of reproduction ©McGraw-Hill Education  Endospores – Figure 3.46 Jump to Endospores – Figure 3.46 Long Description ©McGraw-Hill Education  Eukaryotic Cells – Figure 3.47c Eukaryotic cells are larger and more complex than prokaryotic cells Membrane-enclosed compartments called organelles Nucleus contains cell’s DNA Cells within an organism vary Similar cells form tissues Various tissues form organs ©McGraw-Hill Education © Dr. Thomas Fritsche  Eukaryotic Cells – Figure 3.48 Membrane-bound vesicles bud off from one organelle and fuse with another, delivering material to lumen of organelle A vesicle forms when a The mobile vesicle can then move to section of an organelle other parts of the cell, ultimately fusing buds off. with the membrane of another organelle. ©McGraw-Hill Education  Eukaryotic Cells – Figure 3.47a to b Eukaryotic cells are highly variable Protozoa are self-contained units with no cell wall Animal cells also lack a cell wall Fungal cells have a cell wall containing polysaccharides such as chitin Plant cells have a cell wall composed of cellulose ©McGraw-Hill Education  Eukaryotic Cells – Table 3.6 Table 3.6 Typical Eukaryotic Cell Structures Structure Characteristics Cytoplasmic Membrane Asymmetrical phospholipid bilayer embedded with proteins. Permeability barrier, transport, and cell-to-cell communication. Internal Protein Structures Beat in synchrony to provide movement. Composed of microtubules in a 9 + 2 arrangement. Cilia Cytoskeleton Dynamic filamentous network that provides structure to the cell. Flagella Propel or push the cell with a whip-like or thrashing motion. Composed of microtubules in a 9 + 2 arrangement. Ribosomes Two subunits, 60S and 40S, join to form the 80S ribosome. Membrane-Bound Organelles Site of photosynthesis; the organelle harvests the energy of sunlight to generate ATP, which is then used to Chloroplasts convert CO2 to carbohydrates. Endoplasmic reticulum Site of synthesis of macromolecules destined for other organelles or the external environment. Rough Attached ribosomes thread proteins they are synthesizing into the lumen of the organelle. Smooth Site of lipid synthesis and degradation, and calcium ion storage. Golgi apparatus Site where macromolecules synthesized in the endoplasmic reticulum are modified before being transported in vesicles to other destinations. Lysosome Site of digestion of macromolecules. Mitochondria Harvest the energy released during the degradation of organic compounds to generate ATP. Nucleus Contains the genetic information (DNA). Peroxisome Site where oxidation of lipids and toxic chemicals occurs. ©McGraw-Hill Education  Eukaryotic Cells – Table 3.7 (1) Table 3.7 Comparison of Typical Prokaryotic and Eukaryotic Cell Structures/Functions Prokaryotic Eukaryotic General Characteristics Generally 0.3 to 2 micrometers in diameter. Generally 5 to 50 micrometers in diameter. Size Cell Division Chromosome replication followed by binary fission. Chromosome replication and mitosis followed by division. Chromosome location Located in the nucleoid, which is not membrane- Contained within the membrane-bound nucleus. bound. Structures Relatively symmetrical with respect to the Highly asymmetrical; phospholipid composition of outer layer differs Cytoplasmic membrane phospholipid content of the bilayers. significantly from that of inner layer. Cell wall Composed of peptidoglycan (Bacteria);Gram- Absent in animal cells; composition in other cell types may include negative bacteria have an outer membrane as well. chitin, glucans, and mannans (fungi), and cellulose (plants). Chromosome Single, circular DNA molecule is typical. Multiple, linear DNA molecules. DNA is wrapped around histones. Flagella Composed of protein subunits; attached to the cell Made up of a 9 + 2 arrangement of microtubules; covered by an envelope. extension of the cytoplasmic membrane. Membrane-bound organelles Absent. Present; includes the nucleus, mitochondria, chloroplasts (only in plant cells), endoplasmic reticulum, Golgi apparatus, lysosomes, and peroxisomes. Nucleus Absent; DNA resides as an irregular mass forming Present. the nucleoid region. Ribosomes 70S ribosomes, which are made up of 50S and 30S 80S ribosomes, which are made up of 60S and 40S subunits. subunits Mitochondria and chloroplasts have ribosomes similar to those of bacteria. ©McGraw-Hill Education  Eukaryotic Cells – Table 3.7 (2) Table 3.7 Comparison of Typical Prokaryotic and Eukaryotic Cell Structures/Functions Prokaryotic Eukaryotic Functions Degradation of Enzymes are secreted that degrade Macromolecules can be brought into the cell by endocytosis. extracellular substances macromolecules outside the cell. The resulting Lysosomes carry digestive enzymes. small molecules are transported into the cell. Motility Generally involves flagella, which are composed Involves cilia and flagella, which are made up of a 9 + 2 of protein subunits. Flagella rotate like propellers. arrangement of microtubules. Cilia move in synchrony; flagella propel a cell with a whip-like motion or thrash back and forth to pull a cell forward. Protein secretion Secretion systems transport proteins across the Secreted proteins are moved to the lumen of the rough cytoplasmic membrane. endoplasmic reticulum as they are being synthesized. From there, they are transported to the Golgi apparatus for processing and packaging. Ultimately, vesicles deliver them to the outside of the cell by the process of exocytosis. Strength and rigidity Peptidoglycan-containing cell Cytoskeleton composed of microtubules, intermediate wall (Bacteria). Cytoskeletal components. filaments, and microfilaments. Some have a cell wall; some have sterols in the membrane. Transport Primarily active transport. Group Facilitated diffusion and active transport. Ion channels. translocation (Bacteria). ©McGraw-Hill Education  Cytoplasmic Membrane of Eukaryotic Cells Cytoplasmic membrane similar to prokaryotic cells Phospholipid bilayer embedded with proteins Layer facing cytoplasm differs from that facing outside Proteins in outer layer serve as receptors Bind specific molecule called a ligand Important in cell communication Membranes of many eukaryotes contain sterols Provide strength to otherwise fluid structure Cholesterol in mammals, ergosterol in fungi Lipid rafts: allow cell to detect, respond to signals Many viruses use to enter, exit cells Electrochemical gradient maintained by sodium or proton pumps Membrane not involved in ATP synthesis; performed in mitochondria ©McGraw-Hill Education  Transfer of Molecules Across Cytoplasmic Membrane Aquaporins: water passage Channels: small gated pores, allow small molecules or ions to diffuse Carriers: facilitated diffusion, active transport ©McGraw-Hill Education  Transfer of Molecules Across Cytoplasmic Membrane – Figure 3.49 Endocytosis: cell takes up material from surrounding environment by forming invaginations in cytoplasmic membrane Exocytosis: internal vesicles fuse with the cytoplasmic membrane and release their contents Jump to Transfer of Molecules Across Cytoplasmic Membrane – Figure 3.49 Long Description ©McGraw-Hill Education  Transfer of Molecules Across Cytoplasmic Membrane (1) Pinocytosis most common in animal cells Forms endosome, which fuses to lysosomes Material degraded in endolysosome Receptor-mediated endocytosis allows cell to take up specific extracellular ligands that bind to surface receptors Phagocytosis used by protozoa, phagocytes to engulf Pseudopods surround, bring material into phagosome Phagosome fuses with lysosome to form phagolysosome where material is degraded ©McGraw-Hill Education  Transfer of Molecules Across Cytoplasmic Membrane (2) Secretion Secreted proteins carry a signal sequence that acts as a tag Ribosomes synthesizing protein with a signal sequence attach to endoplasmic reticulum (ER) Protein enters lumen of ER Membrane of ER buds off and binds to cytoplasmic membrane, releasing protein outside the cell Proteins going to other organelles have specific tags ©McGraw-Hill Education  Protein Structures Within Eukaryotic Cells Ribosomes: protein synthesis Eukaryotic ribosome is 80S, made up of 60S and 40S subunits Prokaryotic ribosomes are 70S Antibacterial medications typically do not interfere with 80S ribosome ©McGraw-Hill Education  Protein Structures Within Eukaryotic Cells – Figure 3.50 Cytoskeleton: cell framework Actin filaments (microfilaments) allow movement Polymers of actin polymerize and depolymerize Microtubules are thickest component Long, hollow structures made of tubulin a) Actin filament Found in mitotic spindles, cilia, flagella Framework for organelle and vesicle b) Microtubule movement Intermediate filaments provide mechanical support c) Intermediate filament ©McGraw-Hill Education  Perspective 3.1 Pathogens Hijacking Actin Some pathogens control polymerization and depolymerization of host cell’s actin ©McGraw-Hill Education Top left: © Mark A. Jepson, University of Bristol; Top right: © Pascale F. Cossart; Bottom: © SPL/Science Source  Protein Structures Within Eukaryotic Cells – Figure 3.51 Flagella and cilia function in motility Covered by extensions of cytoplasmic membrane Composed of microtubules in 9 + 2 arrangement Eukaryotic flagella are very different from prokaryotic flagella Propel cell or pull it forward Cilia are shorter than flagella, move synchronously Can move cell forward or move material past stationary cells ©McGraw-Hill Education  Membrane-Bound Organelles – Figure 3.52 Nucleus contains the genetic information Surrounded by two phospholipid bilayer membranes Nuclear pores allow large molecules to pass Nucleolus is region where ribosomal RNAs synthesized ©McGraw-Hill Education b: © Biophoto Associates/Science Source  Membrane-Bound Organelles – Figure 3.53 Mitochondria generate ATP Bounded by two phospholipid bilayers Inner membrane forms folds (cristae), increasing surface area for ATP generation Mitochondrial matrix contains DNA, 70S ribosomes ©McGraw-Hill Education b: © Keith Porter/Science Source  Membrane-Bound Organelles – Figure 3.54 Chloroplasts are site of photosynthesis Found only in plants, algae Harvest sunlight energy to generate ATP ATP used to convert CO2 to sugar and starch Contain DNA and 70S ribosomes, two membranes Stroma contains thylakoids containing pigments that capture radiant energy ©McGraw-Hill Education © Dr. Jeremy Burgess/Science Source  Membrane-Bound Organelles (1) Endosymbiotic theory: ancestors of mitochondria and chloroplasts were bacteria residing within other cells Endosymbiont lost key features (cell wall, replication) as partners because indispensable to one another Lines of evidence Carry DNA for some ribosomal proteins, ribosomal RNA Ribosomes similar to bacterial 70s ribosomes Double membrane surrounds both organelles Multiplication is by binary fission Mitochondrial DNA sequences resemble those of obligate intracellular parasites: rickettsias Chloroplast DNA sequences resemble those of cyanobacteria ©McGraw-Hill Education  Membrane-Bound Organelles – Figure 3.55 Endoplasmic reticulum (ER) System of flattened sheets, sacs, tubes Rough endoplasmic reticulum dotted with ribosomes Synthesize proteins not destined for cytoplasm Smooth endoplasmic reticulum: lipid synthesis and degradation, calcium storage ©McGraw-Hill Education © Don W. Fawcett/Science Source  Membrane-Bound Organelles – Figure 3.56 The Golgi Apparatus Membrane-bound flattened compartments Macromolecules synthesized in ER are modified Addition of carbohydrate, phosphate groups Molecules sorted and delivered in vesicles ©McGraw-Hill Education © Biophoto Associates/Science Source  Membrane-Bound Organelles (2) Lysosomes contain degradative enzymes Could destroy cell if not contained Endosomes, phagosomes fuse with lysosomes Material taken up by cell is degraded Old organelles, vesicles fuse with lysosomes: autophagy Peroxisomes use O2 to degrade lipids, detoxify chemicals Peroxisome generates, contains, and ultimately degrades hydrogen peroxide, superoxide Protects cell from toxic effects of these molecules ©McGraw-Hill Education Appendix of Image Long Descriptions Gram Stain – Figure 3.15a Long Description Steps in Gram stain procedure. Step 1: crystal violet (primary stain) stains all cells purple. Step 2: iodine (mordant) added; all cells remain purple. Step 3: Alcohol (decolorizer) removes color from Gram-negative cells. Gram-positive cells remain purple. Step 4: Safranin (counterstain) makes colorless Gram-negative cells pink. Gram-positive cells remain purple. Jump back to Gram Stain – Figure 3.15a Prokaryotic Cells – Figure 3.24 Long Description Diagram and micrograph showing the positions of structures that make up a prokaryotic cell: capsule, cell wall, cytoplasmic membrane, flagellum, ribosomes, pilus, chromosome, and nucleoid. Jump back to Prokaryotic Cells – Figure 3.24  Permeability of Cytoplasmic Membrane Figure 3.27 (2) Long Description In a hypotonic solution, water flows into the cell; cytoplasmic membrane is pushed against cell wall. In a hypertonic solution, water flows out of the cell; cytoplasmic membrane pulls away from cell wall. Jump back to Permeability of Cytoplasmic Membrane – Figure 3.27 (2)  Transport of Small Molecules Across Cytoplasmic Membrane – Figure 3.31 Long Description The signal sequence on the preprotein targets it for secretion and is removed during the secretion process. Once outside the cell, the protein folds into its functional shape. Extracellular enzymes degrade macromolecules so that the subunits can then be transported into the cell. Jump back to Transport of Small Molecules Across Cytoplasmic Membrane – Figure 3.31 Chemotaxis – Figure 3.40 Long Description A cell moves randomly when there is no concentration gradient of attractant or repellant. When a cell senses it is moving toward an attractant, it tumbles less frequently so the runs in the direction of the attractant are longer. Jump back to Chemotaxis – Figure 3.40 Endospores – Figure 3.46 Long Description Step 1: Cell stops growing; DNA is duplicated. Step 2: A septum forms, dividing the cell asymmetrically. Step 3: The larger compartment engulfs the smaller compartment and the components that will make up the endospore start forming. Step 4: The smaller compartment develops into a forespore, and peptidoglycan-containing material forms between that and the mother cell. Step 5: The mother cell is degraded and the endospore released. It is surrounded by a core wall and cortex, and then by a spore coat. Jump back to Endospores – Figure 3.46 Transfer of Molecules Across Cytoplasmic Membrane – Figure 3.49 Long Description Pinocytosis and receptor-mediated endocytosis form endosomes that fuse with lysosomes to form endolysosomes. Endolysosomes and phagolysosomes form exocytic vesicles that release material across the cytoplasmic membrane. Jump back to Transfer of Molecules Across Cytoplasmic Membrane – Figure 3.49