I. Bacterial Cell Division

1. © 2012 Pearson Education, Inc. I. Bacterial Cell Division • 5.1 Cell Growth and Binary Fission • 5.2 Fts Proteins and Cell Division • 5.3 MreB and Determinants of Cell Morphology • 5.4 Peptidoglycan Synthesis and Cell Division

2. © 2012 Pearson Education, Inc. 5.1 Cell Growth and Binary Fission • Binary fission: cell division following enlargement of a cell to twice its minimum size (Figure 5.1) • Generation time: time required for microbial cells to double in number • During cell division, each daughter cell receives a chromosome and sufficient copies of all other cell constituents to exist as an independent cell

3. Figure 5.1 Cellelongation One generation Septumformation Septum Completionof septum;formation ofwalls; cellseparation © 2012 Pearson Education, Inc.

4. © 2012 Pearson Education, Inc. 5.2 Fts Proteins and Cell Division • Fts (filamentous temperature-sensitive) Proteins (Figure 5.2) • Essential for cell division in all prokaryotes • Interact to form the divisome (cell division apparatus) • FtsZ: forms ring around center of cell; related to tubulin • ZipA: anchor that connects FtsZ ring to cytoplasmic membrane • FtsA: helps connect FtsZ ring to membrane and also recruits other divisome proteins • Related to actin

5. © 2012 Pearson Education, Inc. 5.2Fts Proteins and Cell Division • DNA replicates before the FtsZ ring forms (Figure 5.3) • Location of FtsZ ring is facilitated by Min proteins • MinC, MinD, MinE • FtsK protein mediates separation of chromosomes to daughter cells

6. Figure 5.2 Outer membrane Peptidoglycan Cytoplasmic membrane Divisomecomplex FtsZ ring Cytoplasmic membrane © 2012 Pearson Education, Inc.

7. © 2012 Pearson Education, Inc. 5.3 MreB and Determinants of Cell Morphology • Prokaryotes contain a cell cytoskeleton that is dynamic and multifaceted • MreB: major shape-determining factor in prokaryotes • Forms simple cytoskeleton in Bacteria and probably Archaea • Forms spiral-shaped bands around the inside of the cell, underneath the cytoplasmic membrane (Figure 5.4a and b) • Not found in coccus-shaped bacteria

8. © 2012 Pearson Education, Inc. 5.3 MreB and Determinants of Cell Morphology • MreB (cont’d) • Localizes synthesis of new peptidoglycan and other cell wall components to specific locations along the cylinder of a rod-shaped cell during growth

9. Figure 5.4a FtsZ Cell wall Cytoplasmicmembrane MreB Sites of cellwall synthesis © 2012 Pearson Education, Inc.

10. Figure 5.4b © 2012 Pearson Education, Inc.

11. © 2012 Pearson Education, Inc. 5.3 MreB and Determinants of Cell Morphology • Most archaeal genomes contain FtsZ and MreB-like proteins, thus cell morphology is similar to that seen in Bacteria

12. © 2012 Pearson Education, Inc. 5.4 Peptidoglycan Synthesis and Cell Division • Production of new cell wall material is a major feature of cell division • In cocci, cell walls grow in opposite directions outward from the FtsZ ring • In rod-shaped cells, growth occurs at several points along length of the cell

13. © 2012 Pearson Education, Inc. 5.4 Peptidoglycan Synthesis and Cell Division • Preexisting peptidoglycan needs to be severed to allow newly synthesized peptidoglycan to form • Beginning at the FtsZ ring, small openings in the wall are created by autolysins • New cell wall material is added across the openings • Wall band: junction between new and old peptidoglycan

14. Figure 5.5 FtsZ ring Growth zone Wall bands Septum © 2012 Pearson Education, Inc.

15. Figure 5.4a FtsZ Cell wall Cytoplasmicmembrane MreB Sites of cellwall synthesis © 2012 Pearson Education, Inc.

16. © 2012 Pearson Education, Inc. 5.4 Peptidoglycan Synthesis and Cell Division • Bactoprenol: carrier molecule that plays major role in insertion of peptidoglycan precursors • C55 alcohol (Figure 5.6) • Bonds to N-acetylglucosamine/N-acetylmuramic acid/pentapeptide peptidoglycan precursor

17. © 2012 Pearson Education, Inc. 5.4 Peptidoglycan Synthesis and Cell Division • Glycolases: enzymes that interact with bactoprenol (Figure 5.7a) • Insert cell wall precursors into growing points of cell wall • Catalyze glycosidic bond formation

18. Figure 5.7a Peptidoglycan Growing pointof cell wall Transglycosylaseactivity Autolysinactivity Cytoplasmicmembrane Out In Pentapeptide Bactoprenol © 2012 Pearson Education, Inc.

19. © 2012 Pearson Education, Inc. 5.4 Peptidoglycan Synthesis and Cell Division • Transpeptidation: final step in cell wall synthesis (Figure 5.7b) • Forms the peptide cross-links between muramic acid residues in adjacent glycan chains • Inhibited by the antibiotic penicillin

20. Figure 5.7b Transpeptidation © 2012 Pearson Education, Inc.

21. © 2012 Pearson Education, Inc. II. Population Growth • 5.5 The Concept of Exponential Growth • 5.6 The Mathematics of Exponential Growth • 5.7 The Microbial Growth Cycle

22. © 2012 Pearson Education, Inc. 5.5 The Concept of Exponential Growth • Most bacteria have shorter generation times than eukaryotic microbes • Generation time is dependent on growth medium and incubation conditions

23. © 2012 Pearson Education, Inc. 5.5 The Concept of Exponential Growth • Exponential growth: growth of a microbial population in which cell numbers double within a specific time interval • During exponential growth, the increase in cell number is initially slow but increases at a faster rate (Figure 5.8)

24. © 2012 Pearson Education, Inc. 5.7 The Microbial Growth Cycle • Batch culture: a closed-system microbial culture of fixed volume • Typical growth curve for population of cells grown in a closed system is characterized by four phases (Figure 5.10): • Lag phase • Exponential phase • Stationary phase • Death phase Animation: Bacterial Growth Curve

25. Growth phases Exponential Stationary Death Lag Figure 5.10 1.0 10 0.75 Optical density (OD) 9 Turbidity(optical density) Log10 viableorganisms/ml 0.50 Viable count 8 0.25 7 6 0.1 Time © 2012 Pearson Education, Inc.

26. © 2012 Pearson Education, Inc. 5.7 The Microbial Growth Cycle • Lag phase • Interval between when a culture is inoculated and when growth begins • Exponential phase • Cells in this phase are typically in the healthiest state • Stationary phase • Growth rate of population is zero • Either an essential nutrient is used up or waste product of the organism accumulates in the medium

27. © 2012 Pearson Education, Inc. 5.7 The Microbial Growth Cycle • Death Phase • If incubation continues after cells reach stationary phase, the cells will eventually die

28. © 2012 Pearson Education, Inc. IV. Temperature and Microbial Growth • 5.12 Effect of Temperature on Growth • 5.13 Microbial Life in the Cold • 5.14 Microbial Life at High Temperatures

29. Enzymatic reactions occurringat maximal possible rate Optimum Enzymatic reactions occurringat increasingly rapid rates Figure 5.18 Growth rate Minimum Maximum Temperature Membrane gelling; transportprocesses so slow that growthcannot occur Protein denaturation; collapseof the cytoplasmic membrane;thermal lysis © 2012 Pearson Education, Inc.

30. © 2012 Pearson Education, Inc. 5.12 Effect of Temperature on Growth • Microorganisms can be classified into groups by their growth temperature optima (Figure 5.19) • Psychrophile: low temperature • Mesophile: midrange temperature • Thermophile: high temperature • Hyperthermophile: very high temperature

31. Thermophile Example:Geobacillusstearothermophilus Hyperthermophile Hyperthermophile Figure 5.19 Example:Pyrolobus fumarii Example:Thermococcus celer Mesophile Example:Escherichia coli 60° 106° 88° 39° Growth rate Psychrophile Example:Polaromonas vacuolata 4° 20 30 40 50 60 70 80 90 100 110 120 0 10 Temperature (°C) © 2012 Pearson Education, Inc.

32. © 2012 Pearson Education, Inc. 5.12 Effect of Temperature on Growth • Mesophiles: organismsthat have midrange temperature optima; found in • Warm-blooded animals • Terrestrial and aquatic environments • Temperate and tropical latitudes

33. © 2012 Pearson Education, Inc. 5.13 Microbial Life in the Cold • Extremophiles • Organisms that grow under very hot or very cold conditions • Psychrophiles • Organisms with cold temperature optima • Inhabit permanently cold environments (Figure 5.20) • Psychrotolerant • Organisms that can grow at 0ºC but have optima of 20ºC to 40ºC • More widely distributed in nature than psychrophiles

34. © 2012 Pearson Education, Inc. 5.13 Microbial Life in the Cold • Molecular Adaptations to Psychrophily • Production of enzymes that function optimally in the cold; features that may provide more flexibility • More -helices than -sheets • More polar and less hydrophobic amino acids • Fewer weak bonds • Decreased interactions between protein domains

35. © 2012 Pearson Education, Inc. 5.13 Microbial Life in the Cold • Molecular Adaptations to Psychrophily (cont’d) • Transport processes function optimally at low temperatures • Modified cytoplasmic membranes • High unsaturated fatty acid content

36. Figure 5.22 © 2012 Pearson Education, Inc.

37. Figure 5.23 © 2012 Pearson Education, Inc.

38. © 2012 Pearson Education, Inc. 5.14 Microbial Life at High Temperatures • Studies of thermal habitats have revealed • Prokaryotes are able to grow at higher temperatures than eukaryotes • Organisms with the highest temperature optima are Archaea • Nonphototrophic organisms can grow at higher temperatures than phototrophic organisms

39. © 2012 Pearson Education, Inc. 5.14 Microbial Life at High Temperatures • Molecular Adaptations to Thermophily • Enzyme and proteins function optimally at high temperatures; features that provide thermal stability • Critical amino acid substitutions in a few locations provide more heat-tolerant folds • An increased number of ionic bonds between basic and acidic amino acids resist unfolding in the aqueous cytoplasm • Production of solutes (e.g., di-inositol phophate, diglycerol phosphate) help stabilize proteins

40. © 2012 Pearson Education, Inc. 5.14 Microbial Life at High Temperatures • Molecular Adaptations to Thermophily (cont’d) • Modifications in cytoplasmic membranes to ensure heat stability • Bacteria have lipids rich in saturated fatty acids • Archaea have lipid monolayer rather than bilayer

41. © 2012 Pearson Education, Inc. 5.14 Microbial Life at High Temperatures • Hyperthermophiles produce enzymes widely used in industrial microbiology • Example: Taq polymerase, used to automate the repetitive steps in the polymerase chain reaction (PCR) technique

42. © 2012 Pearson Education, Inc. V. Other Environmental Factors Affecting Growth • 5.15 Acidity and Alkalinity • 5.16 Osmotic Effects on Microbial Growth • 5.17 Oxygen and Microorganisms • 5.18 Toxic Forms of Oxygen

43. © 2012 Pearson Education, Inc. 5.15 Acidity and Alkalinity • The pH of an environment greatly affects microbial growth (Figure 5.24) • Some organisms have evolved to grow best at low or high pH, but most organisms grow best between pH 6 and 8 (neutrophiles)

44. Moles per liter of: pH Example OH H 1 1014 Volcanic soils, watersGastric fluidsLemon juice 101 1013 102 1012 Acid mine drainageVinegar 103 1011 Increasingacidity RhubarbPeaches Acidophiles Figure 5.24 104 1010 Acid soilTomatoes 105 109 American cheeseCabbage 106 108 PeasCorn, salmon, shrimp 107 107 Neutrality Pure water 106 108 Seawater Very alkaline natural soil 105 109 104 1010 Alkaline lakes Increasingalkalinity Soap solutions Alkaliphiles 103 Household ammoniaExtremely alkaline soda lakes 1011 102 1012 Lime (saturated solution) 101 1013 1 1014 © 2012 Pearson Education, Inc.

45. © 2012 Pearson Education, Inc. 5.15 Acidity and Alkalinity • Acidophiles: organisms that grow best at low pH (<6) • Some are obligate acidophiles; membranes destroyed at neutral pH • Stability of cytoplasmic membrane critical • Alkaliphiles: organisms that grow best at high pH (>9) • Some have sodium motive force rather than proton motive force

46. © 2012 Pearson Education, Inc. 5.15 Acidity and Alkalinity • The internal pH of a cell must stay relatively close to neutral even though the external pH is highly acidic or basic • Internal pH has been found to be as low as 4.6 and as high as 9.5 in extreme acido- and alkaliphiles, respectively

47. © 2012 Pearson Education, Inc. 5.15 Acidity and Alkalinity • Microbial culture media typically contain buffers to maintain constant pH

48. © 2012 Pearson Education, Inc. 5.16 Osmotic Effects on Microbial Growth • Typically, the cytoplasm has a higher solute concentration than the surrounding environment, thus the tendency is for water to move into the cell (positive water balance) • When a cell is in an environment with a higher external solute concentration, water will flow out unless the cell has a mechanism to prevent this

49. © 2012 Pearson Education, Inc. 5.16 Osmotic Effects on Microbial Growth • Halophiles: organisms that grow best at reduced water potential; have a specific requirement for NaCl (Figure 5.25) • Extreme halophiles: organisms that require high levels (15–30%) of NaCl for growth • Halotolerant: organisms that can tolerate some reduction in water activity of environment but generally grow best in the absence of the added solute

50. Halophile Halotolerant Extremehalophile Example:Aliivibrio fischeri Example:Staphylococcusaureus Example:Halobacteriumsalinarum Figure 5.25 Growth rate Nonhalophile Example:Escherichia coli 20 10 15 5 0 NaCl (%) © 2012 Pearson Education, Inc.