Prokaryote Cell Structure and Function

1. Prokaryote Cell Structure and Function

2. Background and Classification

3. Prokaryote Cells • No nuclear membrane • No cellular organelles( membrane bound organelles) • Ribosomal size • DNA • RNA • Size • Cell wall and cell membrane

4. A New View of Life • Three Domains of life • Carl Woese responsible for elucidating specific DNA differences between the prokaryotes • Looked at the relationship between the organisms and created a branching tree( see chart)

5. Carl Woese • Studied the molecular biology of the prokaryotes • Used 16s rRNA’s to create his Tree of Life - this is interpreted as an evolutionary distance between types of bacteria in terms of differences in the 16s rRNA • Changes in 16s rRNA may be used as a molecular chronometer or watch to convey the time required to make changes in the genes and proteins – ( Pauling 1965)

6. Parameters used in classification • DNA hybridization – homology of DNA sequences – the use of probes( DNA and m RNA) • G+C content – DNA melting curves. • DNA sequencing • Protein homology • Biochemical characteristics • Molecular characteristics ( expression)

11. Prokaryote Domains Similarities • Bacteria and Archaea have smaller ribosomes ( 70s) • No membrane bound nucleus • Generally one ds circular chromosome- genomic DNA ( there are many exceptions) • Many have plasmids • Operon organization and gene regulation mechanisms Differences • Cell wall differences between Archaea and Eukarya – Peptidoglycan • Cell membane – ester linkage versus ether linkage • Ribosome sensitivity to antibiotics ( chloramphenicol and streptomycin • Ribosomal sensitivity to diptheria toxin • RNA sequences • RNA Polymerases

12. Archaea • Includes organisms regarded a extremophiles • Methanogens • Halophiles • Hyperthermophiles • Nitrogen bacteria

13. Classification - Bacteria • Proteobacteria – Five Classes – largest group. Very diverse • Class I – Alpha proteobacteria – range from Nitrogen fixing bacteria vital to recycling of Nitrogen to pathogens like Rickettsiae • Class II – ( Betaproteobacteria) includes Neisseria species ( gonnorheae and meningitidis ) • Class III( Gammaproteobacteria) includes – E. coli, Salmonella, Shigella, and other pathogens • Class IV – Organisms that are unique – Bdellovibrio that devours gram negative bacteria • Class V – Includes Campylobacter and Helicobacter pylori

14. Gram Positive Bacteria – High G +C content • Actinomyces – Bacteria that are found in the environment • Mycobacterium, actinomyces, and streptomyces • Streptomyces and actinomyces are soil bacteria with unusual characteristics that have contributed to antibiotic therapy ( Selman Waksman – Rutgers)

15. Spirochetes • Unique organisms – • Treponema pallidum • Borrelia • Leptospira

16. Gram Positive Bacteria – Low G-C content • Gram positive organisms • Medically important • Clostridium • Mycoplasma • Bacilli, Enterococcus, and Streptococcus

17. The size of bacteria ranges from 0.1 to about 600 µm over a single dimension They are as small as the largest viruses to large enough for single cells to be visible by the naked eye Prokaryote – Cell Size • Mycoplasmas are about the size of a virus with the diameter of 0.3 µm • E. coli is a more typical bacterium with dimensions of 1.1-1.5 µm wide by 2.0-6.0 µm in length.

18. The range in size • Largest greater than 50 μm in diameter • Smallest less than .3 μm

19. From ultra to nano Epulopiscium fishelsoni Nanobacteria

21. Shapes of bacteria Curved - spirochetes Rods Cocci

22. The Prokaryote Cell

23. Prokaryote Cell Structures

24. Prokaryote Cell Ultrastructure

25. Cell Wall • Rigid structure that lies just outside the plasma membrane • Maintains shape, protects the membrane, and regulates transport

26. Basic Molecular components of the cell wall • Peptidoglycan is a complex polymer of sugars and amino acids • The peptidoglycan that is unique to bacteria is murein. • The fact that murein is unique has made it a target of antibiotics( an entire class) that inhibits the synthesis of the wall. ( Beta lactams which includes penicillin)

27. The basic structure • Glycan sugar chains linked by peptides. • N-acetyl glucosamine • ( NAG) and N- acetyl-muramic acid( NAM) • Linked by four peptide – third is lysine • Cross – Linked with glycines • This structure is similar throughout the Domain bacteria but has variable chemical properties in different species

28. Peptidoglycan • This structure( compared to the chain mail of medieval soldiers) covers the outer surface of the bacterial cell. This determines the shape of the bacterium for instance coccus or bacillus

29. Additional Cell Wall component • An actin like protein has been found underlying bacterial cell walls. • Cytoskeletal elements were previously thought to be absent from bacterial cells • These proteins have been found in gram negative bacteria • This new research indicates that the origin of the eukaryote cell cytoskeleton may be of prokaryote origin.

30. The Two Major Types of Bacterial Cell Walls • Bacteria are divided into two major groups based on the response to Gram-stain procedure. • gram-positive bacteria stain purple • gram-negative bacteria stain pink • staining reaction due to cell wall structure

31. Teichoic Acid • Teichoic acids are found in Gram Positive Cell Walls • Polymers of glycerol or ribitol joined by phosphate groups • Polymers of 30 long • Extend beyond the cell wall

32. Comparison of cell wall structure • The Gram Positive cell wall is characterized by a thick layer of Peptidoglycan. This causes the bacterium to stain purple with the Gram Stain • The Gram Negative cell wall has a layer of lipids overlying the Peptiodglycan layer which is much thinner. This results in a pinkish color upon staining.

33. Gram Stain Technique • Make a smear( spread across the surface of the slide • Air dry smear • Heat fix • Cover smear with Crystal violet – 1 minute • ( gram positive) – purple and rinse • Iodine( mordant) – 1 minute and rinse • Alcohol( decolorizer) – seconds and rinse • Saffranin – gram negative – pink – 1 minute and rinse

34. Gram Staining • Thought to involve constriction of the thick peptidoglycan layer of gram-positive cells • constriction prevents loss of crystal violet during decolorization step • Thinner peptidoglycan layer of gram-negative bacteria does not prevent loss of crystal violet

35. Gram Positive

36. Gram Positive

37. Gram Negative

38. Gram Negative

39. The Outer LPS - Lipopolysaccharide • consist of three parts • lipid A • core polysaccharide • O side chain (O antigen)

41. Characteristics of the Gram Negative Cell Wall • Protection from host defenses (O antigen) • Contributes to negative charge on cell surface (core polysaccharide) • Helps stabilize outer membrane structure (lipid A)

42. LPS • Lipid A is an unusual glycolipid composed of a disaccharide with attached sort-chain fatty acids and phosphate groups. This is linked to fever and shock invertebrates and is an endotoxin

43. LPS • The core –A short series of sugars attached to Lipid A • The O antigen is a long carbohydrate chain up to 40 sugar residues in length which is bound to the core. • The hydrophilic carbohydrate chains of the O antigen exclude hydrophobic compounds

44. Connections • Braun’s lipoproteins connect outer membrane to peptidoglycan • Adhesion sites • sites of direct contact (possibly true membrane fusions) between plasma membrane and outer membrane • substances may move directly into cell through adhesion sites

45. O antigen and importance • The O antigen is highly immunogenic. It elicits a strong antibody response when introduced when introduced into a vertebrate host. • E coli 157:H7 is the pathogenic form of E. coli as compared to a commensal in the gut. This is considered to be a virulence factor.

46. LPS - significance • More permeable than plasma membrane due to presence of porin proteins and transporter proteins • Porin proteins form channels through which small molecules (600-700 daltons) can pass • These proteins and their channels are of great complexity • Larger molecules are translocated by specialized protein complexes

47. Periplasmic space • The two cell wall structures create an internal compartment is the periplasm • This compartment contains degradative enzymes such as nucleases, proteases, and phosphatases • Binding proteins that have a high affinity for amino acids and sugars are also present • It is space that contains the Beta lactamases that degrade antibiotics so that they cannot interfere with the cell wall synthesis

48. Function of LPS and cell wall • Osmotic lysis • Can occur when cells are in hypotonic solutions • Movement of water into cell causes swelling and lysis due to osmotic pressure • Cell wall protects against osmotic lysis

49. Plasmolysis useful in food preservation e.g., dried foods and jellies Osmotic lysis basis of lysozyme and penicillin action Osmotic lysis can occur when cells are in hypotonic solutions movement of water into cell causes swelling and lysis due to osmotic pressure Cell wall protects against osmotic lysis Plasmolysis and Lysis

50. protoplast – the absence ot cell walls in gram-positive spheroplast – the absence of a cell wall in gram-negative