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Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Overview: Microbial Model Systems (This is Ch. 18 in our book, but Ch. 19 according.

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Presentation on theme: "Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Overview: Microbial Model Systems (This is Ch. 18 in our book, but Ch. 19 according."— Presentation transcript:

1 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Overview: Microbial Model Systems (This is Ch. 18 in our book, but Ch. 19 according to F & T Guide.) Viruses called bacteriophages can infect and set in motion a genetic takeover of bacteria, such as Escherichia coli E. coli and its viruses are called model systems because of their frequent use by researchers in studies that reveal broad biological principles Beyond their value as model systems, viruses and bacteria have unique genetic mechanisms that are interesting in their own right

2 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings What’s going on here?

3 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Bacteria are prokaryotes w ith cells much smaller and more simply organized than those of eukaryotes Viruses a re smaller and simpler than bacteria

4 Comparison of animal cell, bacterial cell, and virus Virus Bacterium Animal cell Animal cell nucleus 0.25 µm

5 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 1. Concept 18.1: A virus has a genome but can reproduce only within a host cell Scientists detected viruses indirectly long before they could see them The story of how viruses were discovered begins in the late 1800s

6 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 1—2. The Discovery of Viruses: Scientific Inquiry Tobacco mosaic disease stunts growth of tobacco plants and gives their leaves a mosaic coloration In the late 1800s, researchers hypothesized that a particle smaller than bacteria caused the disease 1883—Adolf Mayer 1893—Dimitri Ivanowsky In 1935, Wendell Stanley confirmed this hypothesis by crystallizing the infectious particle, now known as tobacco mosaic virus (TMV)

7 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Leaf on right is infected with TMV

8 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Structure of Viruses Viruses are not cells Viruses are very small infectious particles consisting of nucleic acid enclosed in a protein coat and, in some cases, a membranous envelope

9 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 3. Viral Genomes Viral genomes may consist of – Double- stranded DNA – Single-stranded DNA – Double- stranded RNA – Single-stranded RNA Depending on its type of nucleic acid, a virus is called a DNA virus or an RNA virus

10 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 4. Capsids and Envelopes A capsid i s the protein shell that encloses the viral genome Capsomeres are the protein subunits that make up the capsid A capsid can have various structures

11 4. Helical capsid Capsomere of capsid RNA 18  250 mm Tobacco mosaic virus 20 nm

12 4. Polyhedral capsid Capsomere Glycoprotein 70–90 nm (diameter) DNA Adenoviruses 50 nm

13 4. Polyhedral head and tail apparatus 80  225 nm DNA Head Tail sheath Tail fiber Bacteriophage T4 50 nm

14 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Some viral structures have membranous envelopes that help them infect hosts These viral envelopes surround the capsids of influenza viruses and many other viruses found in animals Viral envelopes, which are derived from the host cell’s membrane, contain a combination of viral and host cell molecules They contain host phospholipids and membrane proteins and viral proteins and glycoproteins 5. Viral envelopes

15 5. Enveloped virus Glycoprotein 80–200 nm (diameter) RNA Capsid Influenza viruses 50 nm Membranous envelope

16 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 6. Role of the viral envelope An animal virus with a viral envelope uses it to enter the host cell Viral glycoproteins bind to specific receptor molecules on the surface of a host cell

17 6. RNA ER Capsid HOST CELL Viral genome (RNA) mRNA Capsid proteins Envelope (with glycoproteins) Glyco- proteins Copy of genome (RNA) Capsid and viral genome enter cell New virus Template

18 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Bacteriophages, also called phages, are viruses that infect bacteria They have the most complex capsids found among viruses Phages have an elongated capsid head that encloses their DNA A protein tailpiece attaches the phage to the host and injects the phage DNA inside 7.

19 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 8—9. General Features of Viral Reproductive Cycles Viruses identify their host cells by a “lock-and-key” fit between proteins on the outside of the virus and specific receptor molecules on the surface of cells Viruses are obligate intracellular parasites, which means they can reproduce only within a host cell Animation: Simplified Viral Reproductive Cycle Animation: Simplified Viral Reproductive Cycle

20 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 10—11. Each virus has a host range, a limited number of host cells that it can infect Broad host range: West Nile virus (mosquitoes, birds, and humans) Narrow host range: measles virus and poliovirus can only infect humans Rabies virus has a broad host range (mammals such as dogs, raccoons, bats, humans), but a human cold virus can only infect humans

21 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 12. Viruses use enzymes, ribosomes, and small host molecules (nucleotides, amino acids) to synthesize progeny viruses

22 12, 13, 15. DNA VIRUS Capsid HOST CELL Viral DNA Replication Entry into cell and uncoating of DNA Transcription Viral DNA mRNA Capsid proteins Self-assembly of new virus particles and their exit from cell

23 14. RNA ER Capsid HOST CELL Viral genome (RNA) mRNA Capsid proteins Envelope (with glycoproteins) Glyco- proteins Copy of genome (RNA) Capsid and viral genome enter cell New virus Template

24 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Reproductive Cycles of Phages Phages are the best understood of all viruses Phages have two reproductive mechanisms: the lytic cycle and the lysogenic cycle

25 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 16, 18. The Lytic Cycle The lytic cycle is a phage reproductive cycle that culminates in the death of the host cell The lytic cycle produces new phages and digests the host’s cell wall, releasing the progeny viruses A phage that reproduces only by the lytic cycle is called a virulent phage Bacteria have defenses against phages, including restriction enzymes that recognize and cut up certain phage DNA

26 The Lytic Cycle Attachment Entry of phage DNA and degradation of host DNA Synthesis of viral genomes and proteins Assembly Release Phage assembly Head Tails Tail fibers

27 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 17. The _______ of the phage enters the cell, leaving the empty _________ outside. This is accomplished when the sheath of the __________ contracts.

28 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 18, 19, 20. Bacterial defenses against phages: 1. Natural selection favors bacterial mutants with receptor sites that are no longer recognized by phages. 2. As soon as phage DNA enters the bacterial host cell, it fights back with restriction enzymes, which hydrolyze the viral DNA. (The bacterial cell’s own DNA is chemically modified to protect it from its own restriction enzymes.) 3. Not all phages kill their hosts—some use the lysogenic cycle

29 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 16, 21. The Lysogenic Cycle The lysogenic cycle replicates the phage genome without destroying the host The viral DNA molecule is incorporated by genetic recombination into the host cell’s chromosome This integrated viral DNA is known as a prophage Every time the host divides, it copies the phage DNA and passes the copies to daughter cells Phages that use both the lytic and lysogenic cycles are called temperate phages Animation: Phage Lambda Lysogenic and Lytic Cycles Animation: Phage Lambda Lysogenic and Lytic Cycles

30 23, 24. Phage DNA The phage attaches to a host cell and injects its DNA. Phage DNA circularizes Bacterial chromosome Lytic cycle The cell lyses, releasing phages. Lytic cycle is induced or Lysogenic cycle is entered Certain factors determine whether Lysogenic cycle Occasionally, a prophage exits the bacterial chromosome, initiating a lytic cycle. The bacterium reproduces normally, copying the prophage and transmitting it to daughter cells. Prophage Many cell divisions produce a large population of bacteria infected with the prophage. Daughter cell with prophage Phage DNA integrates into the bacterial chromosomes, becoming a prophage. New phage DNA and proteins are synthesized and assembled into phages.

31 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 22. The Trojan horse was full of Greek soldiers who, once they were within the walls of Troy, leapt out and destroyed the city. Prophages “hide out” within the host cell and are replicated along with the host’s DNA. They might be triggered to “go lytic” by an environmental signal, such as radiation or the presence of certain chemicals.

32 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 25. Few bacteriophages have an envelope or RNA genome, but… Animal viruses often have an envelope derived from the host cell’s membrane or nuclear envelope Animal viruses often have an RNA genome

33 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Classifying Animal Viruses Two key variables in classifying viruses that infect animals: – DNA or RNA? – Single-stranded or double-stranded?

34 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Class/FamilyEnvelopeExamples/Disease I. Double-stranded DNA (dsDNA) AdenovirusNo Respiratory diseases, animal tumors PapovavirusNo Papillomavirus (warts, cervical cancer): polyomavirus (animal tumors) HerpesvirusYes Herpes simplex I and II (cold sores, genital sores); varicella zoster (shingles, chicken pox); Epstein-Barr virus (mononucleosis, Burkitt’s lymphoma) PoxvirusYes Smallpox virus, cowpox virus

35 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Class/FamilyEnvelopeExamples/Disease II. Single-stranded DNA (ssDNA) ParvovirusNoB19 parvovirus (mild rash) III. Double-stranded RNA (dsRNA) ReovirusNoRotavirus (diarrhea), Colorado tick fever virus

36 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Class/FamilyEnvelopeExamples/Disease IV. Single-stranded RNA (ssRNA); serves as mRNA PicornavirusNoRhinovirus (common cold); poliovirus, hepatitis A virus, and other enteric (intestinal) viruses CoronavirusYesSevere acute respiratory syndrome (SARS) FlavivirusYesYellow fever virus, West Nile virus, hepatitis C virus TogavirusYesRubella virus, equine encephalitis viruses

37 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Class/FamilyEnvelopeExamples/Disease V. ssRNA; template for mRNA synthesis FilovirusYes Ebola virus (hemorrhagic fever) OrthomyxovirusYes Influenza virus ParamyxovirusYes Measles virus; mumps virus RhabdovirusYes Rabies virus VI. ssRNA; template for DNA synthesis RetrovirusYes HIV (AIDS); RNA tumor viruses (leukemia)

38 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 26. RNA as Viral Genetic Material The broadest variety of RNA genomes is found in viruses that infect animals Retroviruses use reverse transcriptase to copy their RNA genome into DNA HIV is the retrovirus that causes AIDS

39 27. Capsid Viral envelope Glycoprotein Reverse transcriptase RNA (two identical strands)

40 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The viral DNA that is integrated into the host genome is called a provirus Unlike a prophage, a provirus remains a permanent resident of the host cell The host’s RNA polymerase transcribes the proviral DNA into RNA molecules The RNA molecules function both as mRNA for synthesis of viral proteins and as genomes for new virus particles released from the cell 28.

41 29. HOST CELL Reverse transcription Viral RNA RNA-DNA hybrid DNA NUCLEUS Chromosomal DNA Provirus RNA genome for the next viral generation mRNA New HIV leaving a cell HIV entering a cell 0.25 µm HIV Membrane of white blood cell

42 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 30. Evolution of Viruses Viruses do not fit our definition of living organisms Since viruses can reproduce only within cells, they probably evolved from bits of cellular nucleic acid, such as… Plasmids, which are small, circular DNA molecules found in bacteria and yeast, or Transposons, which are DNA segments that can move around within a cell’s genome.

43 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Concept 18.2: Viruses, viroids, and prions are formidable pathogens in animals and plants Diseases caused by viral infections affect humans, agricultural crops, and livestock worldwide Smaller, less complex entities called viroids and prions also cause disease in plants and animals

44 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 31. Viral Diseases in Animals Viruses may damage or kill cells by causing the release of hydrolytic enzymes from lysosomes Some viruses cause infected cells to produce toxins that lead to disease symptoms Some have molecular components that are toxic, such as membrane proteins Cold viruses attack epithelial cells, which easily replace themselves through cell division. Poliovirus attacks nerve cells, which are irreparable.

45 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Our immune system provides natural defense against viruses Modern medicine has developed vaccines and drugs Vaccines are harmless derivatives of pathogenic microbes that stimulate the immune system to mount defenses against the actual pathogen Vaccines can prevent certain viral illnesses Antiviral drugs include -acyclovir, which interferes with viral nucleic acid synthesis -AZT, which interferes with reverse transcriptase 32. Fighting back against viruses:

46 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 33. Emerging Viruses Emerging viruses are those that appear suddenly or suddenly come to the attention of scientists Severe acute respiratory syndrome (SARS) recently appeared in China Outbreaks of “new” viral diseases in humans are usually caused by: --Mutation of existing viruses creating new strains (ex. flu) --Spread of existing viruses from one host species to another (ex. hantavirus, bird flu, swine flu) --Cultural change (ex. people traveling more, blood transfusions, IV drug use, and sexual promiscuity all contributed to the spread of HIV)

47 33. Young ballet students in Hong Kong wear face masks to protect themselves from the virus causing SARS. The SARS-causing agent is a coronavirus like this one (colorized TEM), so named for the “corona” of glyco-protein spikes protruding form the envelope.

48 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 34. H1N1 H and N stand for the two glycoproteins that are on the surface of the viral envelope of this flu virus (their scientific names are hemagglutinin and neuramidinase) and do its dirty work. There are 16 types of the H protein, numbered H1 through H16, and 9 types of the N protein, numbered N1 through N9. That makes 144 possible combinations of the virus, a constantly changing challenge for prevention or treatment. A new combination, H2N2, cause a brief swine flu epidemic in An H3N2 strain was the source of another epidemic in The bird flu virus that began in Southeast Asia a decade ago and has spread throughout the Old World is an H5N1 combination.

49 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Viral Diseases in Plants More than 2,000 types of viral diseases of plants are known Some symptoms are spots on leaves and fruits, stunted growth, and damaged flowers or roots

50 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings

51 Plant viruses spread disease in two major modes: – Horizontal transmission, entering through damaged cell walls – Vertical transmission, inheriting the virus from a parent 35.

52 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 36. Once inside a plant cell, the virus replicates and can spread throughout the plant body through plasmodesmata Some viral genes code for proteins that increase the diameter of plasmodesmata so the viruses can spread more easily!

53 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Viroids and Prions: The Simplest Infectious Agents Viroids are circular RNA molecules that infect plants and disrupt their growth They teach us that even a single molecule can be an infectious agent that spreads disease One viroid disease killed 10 million coconut palms in the Philippines Prions are slow-acting, virtually indestructible infectious proteins that cause brain diseases in mammals Prions propagate by converting normal proteins into the prion version Prions are transmitted in food

54 38. Prions propagating Normal protein New prion Prion Original prion Many prions

55 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 39. Diseases caused by prions Scrapie (in sheep) Mad cow disease Creutzfeldt-Jakob disease Kuru (laughing disease of the Fore tribe of Papau, New Guinea)

56 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 40, 41. Prions are scary The diseases they cause are all neurodegenerative The diseases they cause are incurable Stanley B Prusiner (in 1982) claimed to have isolated prions (he also named them) and won the Nobel prize in 1997

57 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Concept 18.3: Rapid reproduction, mutation, and genetic recombination contribute to the genetic diversity of bacteria Now we’re working on part of a F & T Guide (it’s their Chapter 27, but still our Chapter 18)! Sorry for the confusion!! Bacteria allow researchers to investigate molecular genetics in the simplest true organisms The well-studied intestinal bacterium Escherichia coli (E. coli) is “the laboratory rat of molecular biology”

58 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 14. The Bacterial Genome and Its Replication The bacterial chromosome is usually a circular DNA molecule with few associated proteins Many bacteria also have plasmids, smaller circular DNA molecules that can replicate independently of the chromosome Bacterial cells divide by binary fission, which is preceded by replication of the chromosome

59 Not in the Lecture Guide, but you need to know… Origin of replication Replication fork Termination of replication

60 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 15, 16. See book, page 536. Endospores are dormant, non-reproductive forms of bacteria. They are formed due to environmental factors, such as when nutrient levels are low. Bacteria can survive in this dormant state for centuries. Ex: Bacillus anthracis (anthrax), Clostridium tetani (tetanus)

61 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 17. Mechanisms of Gene Transfer and Genetic Recombination in Bacteria Three processes bring bacterial DNA from different individuals together: – Transformation – Transduction – Conjugation

62 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 17, 18. Transformation Transformation is the alteration of a bacterial cell’s genotype and phenotype by the uptake of naked, foreign DNA from the surrounding environment For example, harmless Streptococcus pneumoniae bacteria can be transformed to pneumonia-causing cells (Frederick Griffith’s experiment)

63 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 17, 19, 20. Transduction In the process known as transduction, phages carry bacterial genes from one host cell to another

64 Generalized Transduction A+A+ Phage DNA A+A+ Donor cell B+B+ A+A+ B+B+ Crossing over A+A+ A–A– B–B– Recipient cell A+A+ B–B– Recombinant cell

65 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Conjugation and Plasmids Conjugation is the direct transfer of genetic material between bacterial cells that are temporarily joined The transfer is one-way: One cell (“male”) donates DNA, and its “mate” (“female”) receives the genes

66 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings “Maleness,” the ability to form a sex pilus and donate DNA, results from an F (for fertility) factor as part of the chromosome or as a plasmid Plasmids, including the F plasmid, are small, circular, self-replicating DNA molecules 21.

67 21, 22. Sex pilus 5 µm

68 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 22, 23, 24. The F Plasmid and Conjugation Cells containing the F plasmid, designated F + cells, function as DNA donors during conjugation F + cells transfer DNA to an F  recipient cell Chromosomal genes can be transferred during conjugation when the donor cell’s F factor is integrated into the chromosome A cell with a built-in F factor is called an Hfr cell The F factor of an Hfr cell brings some chromosomal DNA along when transferred to an F – cell

69 24. Conjugation F plasmidBacterial chromosome F + cell Mating bridge F + cell Bacterial chromosome F – cell Conjunction and transfer of an F plasmid from and F + donor to an F – recipient

70 22, 25, 26. Episomes and Hfr cells F plasmidBacterial chromosome F + cell Mating bridge F + cell Bacterial chromosome F – cell Conjunction and transfer of an F plasmid from and F + donor to an F – recipient. The F Factor is incorporated into the host’s chromosome and is called an episome F + cell Hfr cell F factor

71 27. Hfr stands for “high frequency of recombination, because when these cells conjugate with other cells, there’s a good chance genetic recombination is going to occur: F plasmidBacterial chromosome F + cell Mating bridge F + cell Bacterial chromosome F – cell Conjunction and transfer of an F plasmid from and F + donor to an F – recipient F + cell Hfr cell F factor Hfr cell F – cell

72 27. F plasmid Bacterial chromosome F + cell Mating bridge F + cell Bacterial chromosome F – cell Conjunction and transfer of an F plasmid from and F + donor to an F – recipient F + cell Hfr cell F factor Hfr cell F – cell Temporary partial diploid Recombinant F – bacterium Conjugation and transfer of part of the bacterial chromosome from an Hfr donor to an F – recipient, resulting in recombiination

73 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 28. R plasmids and Antibiotic Resistance R plasmids confer resistance to various antibiotics When a bacterial population is exposed to an antibiotic, individuals with the R plasmid will survive and increase in the overall population

74 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 29. Transposition of Genetic Elements Transposable elements are sections of DNA that can move around with a cell’s genome The DNA of a cell can also undergo recombination due to the movement of transposable elements Transposable elements, often called “jumping genes,” contribute to genetic shuffling in bacteria

75 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 30. Insertion Sequences Insertion sequences are the simplest transposable elements and exist only in bacteria An insertion sequence has a single gene for transposase, an enzyme catalyzing movement of the insertion sequence from one site to another within the genome

76 30. An insertion sequence Insertion sequence Transposase gene 5 3 Inverted repeat 3 5 Inverted repeat

77 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 31. Transposons Transposable elements called transposons are longer and more complex than insertion sequences In addition to DNA required for transposition, transposons have extra genes that “go along for the ride,” such as genes for antibiotic resistance

78 31. Transposons Transposon Insertion sequence Insertion sequence Antibiotic resistance gene Transposase gene Inverted repeat

79 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 1, 2. Concept 18.4: Individual bacteria respond to environmental change by regulating their gene expression Now, we’re continuing on to the Chapter 18 F & T Guide! (still our Chapter 18!) A bacterium can tune its metabolism to the changing environment and food sources This metabolic control occurs on two levels: – Adjusting activity of metabolic enzymes – Regulating genes that encode metabolic enzymes

80 2, 3. Regulation of enzyme activity Regulation of enzyme production Enzyme 1 Regulation of gene expression Enzyme 2 Enzyme 3 Enzyme 4 Enzyme 5 Gene 2 Gene 1 Gene 3 Gene 4 Gene 5 Tryptophan Precursor Feedback inhibition

81 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 4—7. Regulation of Gene Expression in Prokaryotes Involves Operons: In bacteria, genes are often clustered into operons, composed of – An operator, an “on-off” switch – A promoter, the place where _______ binds – Genes for metabolic enzymes An operon can be switched off by a protein called a repressor A corepressor is a small molecule that cooperates with a repressor to switch an operon off

82 Example: the trp operon, a repressible operon Promoter DNA trpR Regulatory gene RNA polymerase mRNA 3 5 Protein Inactive repressor Tryptophan absent, repressor inactive, operon on mRNA 5 trpE trpD trpC trpBtrpA Operator Start codon Stop codon trp operon Genes of operon E Polypeptides that make up enzymes for tryptophan synthesis D C B A

83 The repressor protein is coded for by a regulatory gene (not part of the operon) in an inactive form: DNA Protein Tryptophan (corepressor) Tryptophan present, repressor active, operon off mRNA Active repressor The repressor is activated when the corepressor, tryptophan, is present

84 The activated repressor binds to the operator, which blocks RNA polymerase from binding. The gene is turned __________. DNA Protein Tryptophan (corepressor) Tryptophan present, repressor active, operon off mRNA Active repressor No RNA made

85 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 8—9. Repressor proteins bind to the operator and block RNA polymerase from binding to the promoter. This STOPS transcription The regulatory gene codes for the repressor protein. The regulatory gene is NOT part of the operon and is usually located elsewhere on the bacterial chromosome

86 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 10. Repressible and Inducible Operons: Two Types of Negative Gene Regulation A repressible operon is one that is usually on; binding of a repressor to the operator shuts off transcription The trp operon is a repressible operon An inducible operon is one that is usually off; a molecule called an inducer inactivates the repressor and turns on transcription The classic example of an inducible operon is the lac operon, which contains genes coding for enzymes in hydrolysis and metabolism of lactose

87 The lac operon. The repressor is coded for by the regulatory gene in an active form. DNA lacl Regulatory gene mRNA 5 3 RNA polymerase Protein Active repressor No RNA made lacZ Promoter Operator Lactose absent, repressor active, operon off

88 11. DNAlacl mRNA 5 3 lac operon Lactose present, repressor inactive, operon on lacZ lacYlacA RNA polymerase mRNA 5 Protein Allolactose (inducer) Inactive repressor  -Galactosidase Permease Transacetylase

89 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Inducible enzymes usually function in catabolic pathways. Their repressors are made in an ____________ form and require an __________ before they can turn the gene _________. Repressible enzymes usually function in anabolic pathways. Their repressors are made in an ____________ form and require a __________ before they can turn the gene _________. Regulation of the BOTH trp and lac operons involves negative control of genes because, ultimately, operons are switched off by the active form of the repressor 12.

90 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 13, 17. Negative regulation Remember, both repressible and inducible operons are eventually turned OFF when the repressor protein binds. Therefore, they are both examples of negative control.

91 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 14. Positive Gene Regulation Some operons are also subject to positive control through a stimulatory activator protein, such as catabolite activator protein (CAP) When glucose (a preferred food source of E. coli ) is scarce, the lac operon is activated by the binding of CAP When glucose levels increase, CAP detaches from the lac operon, turning it off

92 14. DNA cAMP lacl CAP-binding site Promoter Active CAP Inactive CAP RNA polymerase can bind and transcribe Operator lacZ Inactive lac repressor Lactose present, glucose scarce (cAMP level high): abundant lac mRNA synthesized

93 14. DNA lacl CAP-binding site Promoter RNA polymerase Is lazy. Operator lacZ Inactive lac repressor Inactive CAP Lactose present, glucose present (cAMP level low): little lac mRNA synthesized

94 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 15. Positive regulation CAP is an example of positive regulation of a gene When CAP is bound to the CAP-binding site, it increases RNA polymerase’s affinity for the promoter, and increases transcription of the genes You can think of the repressor protein as an on-off switch and CAP as volume control

95 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 16. When glucose levels are high, cAMP levels will be _____________ and active CAP levels will be _____________. When glucose levels are low, cAMP levels will be _____________ and active CAP levels will be _____________.

96 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings FINALLY! We’re done with OUR Chapter 18 and we’ve covered all the topics in that Chapter. Now, on to Chapter 19, even though we’re still working on the Ch. 18 F & T Guide… :\


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