Medical Microbiology 02

8/3/2019 Medical Microbiology 02

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Chapter 3

Bacterial Metabolism and Genetics


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Bacterial Metabolism

Metabolic requirements

Metabolism, Energy, and Biosynthesis

Metabolism of Glucose

Embden-Meyerhof-Parnas Pathway

Tricarboxylic Acid Cycle

Pentose Phosphate Pathway


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Metabolic Requirements

Minimum requirement for growth

Oxygen requirements

Classification based on nutritional requirements


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Minimum requirements for growth

C, O, H, N, S, P

K, Na, Mg, Ca, Cl

Fe, Zn, Mn, Mo, Se, Co, Cu, Ni

Many bacteria secrete special proteins,siderophores, to concentrate iron from dilutesolutions.


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Oxygen requirements

Obligate anaerobes: Clostridium perfringens

Obligate aerobes: Mycobacterium tuberculosis

Facultativeanaerobes: Escherichia coli


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Classification based on nutritional requirements

autotrophs

lithotrophs

organotrophs


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Metabolism, Energy, and Biosynthesis

catabolism: substrate breakdown and conversion

anabolism: synthesis of cellular constituentsThe metabolites are converted via one or morepathways into pyruvate.

From pyruvate, carbons are channeled to energyproduction or the synthesis of new carbohydrates,amino acids, lipids and nucleic acids.


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Metabolism of Glucose

production of energy of useful forms

production of energy through fermentation,anaerobic respiration, or aerobic respiration


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Embden-Meyerhof-Parnas Pathway (Glycolysis)

Converting glucose to pyruvate

substrate level phosphorylation:generating ATP from ADP via

kinases

conversion of NAD to NADHproduces ATP (via electrontransport chain)

Fermentation starts frompyruvate.


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Fermentation

Organic molecules, ratherthan oxygen, are used aselectron acceptors torecycle NADH.

Monosodium glutamate isthe largest fermentationproduct in the world.


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Tricarboxylic Acid Cycle

Pyruvate is completelyoxidized to water and CO2 inthe presence of O2 via theTCA cycle.

Oxaloacetate and acetyl-CoAcondense to form citrate.

Citrate is then converted tooxaloacetate through aseries of steps.

Produces 2 CO2, 3 NADH, 1FADH2, and 1 GTP.


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Electron Transport Chain

The electrons carried byNADH are passed through a

stepwise fashion through aseries of donor-acceptorpairs and ultimately to O2(aerobic respiration) or otherterminal acceptors, includingnitrate, sulfate, CO2, ferric

ion (anaerobic respiration).


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Energy Generated throughAerobic Glucose Metabolism


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Importance of the TCA Cycle

the most efficientmechanism for generatingATP

the final common pathway

for the complete oxidationof amino acids, fatty acids,and carbohydrates

supplying keyintermediates, e.g. -ketoglutarate, pyruvate,oxaloacetate for theultimate synthesis ofamino acids, lipids,purines, and pyrimidines


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Pentose Phosphate Pathwayor Hexose monophosphate

Shunt

Ribulose-5-phosphate isconverted to ribose-5-phosphate, a precursor ofnucleotide biosynthesis,

or xylulose-5-phosphate.

The pathway usestransketolases andtransaldolases togenerate various sugars,

which are usedmetabolically or shunt intothe glycolysis pathway.


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The Bacterial Genes

and Expression

Transcription

Translation

Control of Gene Expression


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Transcription

mRNA

DNA-dependent RNApolymerase

sigma factors

promoters and operators

operons

polycistronic mRNA


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Template and Sense Strands

CAAGAAAAAT AAATTAATTA ATACTATATT CAAAAAAACT GTTATAAAAA

TTTTATATTT TTTATTGTAC TTTAAAAAAA TTCGGTTA AT ATAATGCATA

TATGGATTAT ATAGTCATAT AATTTCTTTT TATTGTAATT ATTTCAGTTT

CTTATCCTCT TATAAATTAG AATTGGAGGG AATTCGTTGA AAGAAAATAC

-51

-1

+50

+100

M K E N TRBS

+1

-10-35


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Translation -- tRNA andmRNA

tRNA

codon

anticodon


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Translation -- Reading Frames


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Translation -- Initiation and TerminationCodons

initiation codons: AUG, GUG, UUG

termination (nonsense) codons: UAA (ochre), UAG (amber), UGA

(opal)

nonsense mutations: a sense codon that is mutated to a nonsensecodon

nonsense suppressor mutation (a conditional lethal mutation) -- a tRNA

mutation at the anti-codon loop that allows readthrough of a nonsensemutation.


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Translation -- Initiation

How do bacteria choose an AUGcodon to initiate translation?

What is the function of ribosomal-binding site (RBS)?

RBS is an A-G-rich sequence, 4-n.t. long, located about 10

nucleotides upstream of aninitiation codon.

RBS complements the 3 region of

16S rRNA in the ribosome.

What is the amino acid that startsthe sequence of a protein whenGUG (Val) or UUG (Leu) is usedfor initiation?

Transpeptidation


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Control of Gene Expression

sigma factor and gene expression

quorum sensing: N-acyl homoserine lactone (AHL) or small peptides

cAMP

pathogenicity island

negative control: repressors

operators

positive control: apoinducer


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The lactose operon

E. colidoes not use lactose whenglucose is present.

Allolactose is the inducer.

RNA polymerase does not have astrong affinity to the lacpromoter.

LacI, the repressor, promotes thebinding of RNA polymerase to thepromoter.

The CAP-cAMP complex promotesthe binding of RNA polymerase,through the C-terminal domain of the -subunit.


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The Tryptophan Operon

Repression of the operon requires TrpR.

Tryptophan is a corepressor.

Binding of TrpR to the operator blocks thebinding of RNA polymerase to thepromoter.

The expression of the operon depends onthe translation of the leader region of themRNA (posttranscriptional control;attenuation).


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helicase, primase gyrase,DNA polymerases, DNA

ligase

DNA cannot be synthesizedwithout a primer.

semiconservative synthesis

bidirectional synthesis

DNA Replication


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Bacterial Growth


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Population Dynamics


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Mutations

transition, transversion

silent mutation, missense mutation, nonsense mutation,frameshift mutations, null mutation, insertional mutation, deletion,inversion

conditional lethal mutations: temperature-sensitive mutations,suppressor mutations

reversion

mutagens

Ames test


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Carcinogenicity Test (Ames Test)

Do you know why ahistidine mutant instead ofa wild-type Salmonellastrain is used in Ames test?


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DNA Repair

direct DNA repair

excision repair

recombinational or postreplicational repair

the SOS response

error-prone repair


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Gene Exchange in Prokaryotic cells

plasmids

bacteriophages

fertility factor

transposons


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Plasmids

circular or linear

containing a replicon

containing a constant copy number

carrying genes that are necessary for plasmid replication

may contain genes critical to bacterial survival

F, the fertility factor, is involved in conjugation


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Bacteriophages

T

4

T

5

T

7

P

2

P22


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Transposons

IS10, Tn10

transposases: encoded by tnpor tnpA;enzyme necessary for transposition

IS: insertion sequences; insertionsequences in a composite transcription

are the same type

nonreplicative (cut-and-paste) andreplicative transposition

tnpR, the resolvase gene; necessary for

the resolution of the cointegrate form bythe Tn3-(TnA) family of transposons;Res site: resolution site or binding site forTnpR, the resolvase

Mu and HIV are actually transposons.


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Replicative Transposition (TnA, Tn3)

Resolvase


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Noneplicative Transposition


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Mechanisms of Genetic Transfer

transformation

conjugation

transduction

transposition


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Transformation

Not all the bacterial speciescan be transformed.


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Conjugation


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Transduction

generalized transduction

specialized transduction


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Recombination

homologous (legitimate) recombination

nonhomologous (illegitimate) recombination

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Medical Microbiology 02