Chapter 7

DNA (Deoxyribonucleic Acid)

Genome-complete set of genetic information

  • All cells have DMA.

  • in bacteria -includes plasmids

  • Gene-functional unit of the genome

Genetics - Study of the function and Transfer of genes

Genomics - Study and analysis of DNA nucleotide Sequence

History

1866 - Gregor Mendel determined traits are invited as physical units

  • Gens

1941 - Beadle and Tatum - genes drive the production of enzymes

  • Treated mold

  • metabolic defect inherited as a single gene

  • one gene-one enzyme

  • won Nobel prize

DNA Replication

A copy of DNA is created for cell division to occur

The bacterial genome is a circle

  • replication is different than eukaryotic

bidirectional - half the time

semiconservative - one old strand, one new strand

lots of enzymes are involved

DNA polymerase creates a new strand of DNA using an original strand as a template

Prokaryotes have an origin of replication

Hydrolysis of a phosphate group from dATP, dTTP, and dGTP provides energy

New nucleotide always added to the 3' end

one mistake per billion bases

40 min for E.Coli to replicate its genome

  • replication can begin before one ends

Each daughter cell inherits one complete chromosome already undergoing replication

Gene Expression

As transcription stops, mRNA rapidly degrade and levels drop

Gene expression can be controlled

Transcription

RNA Polymerase binds to a promoter region of the

DNA and creates a single-stranded complementary RNA (mRNA) in the 5' to 3' direction

In bacteria, mRNA can contain the blueprints for one gene (monocistronic) or many genes (poly)

  • Polycistronic messages usually encode for a group of proteins in the same metabolic pathway

    • Allows for regulation schemes

Plus Strand - coding strand

minus Strand - Template strand

Promoter is upstream of the gene

  • no primer needed

Transcription ends at a terminator

Initiation

RNA polymerase complex recognizes a promoter region on the dsDNA

The sigma factor subunit does the recognizing and dissociates leaving the rest of the enzyme to complete the elongation

  • Different from eukaryotic and arches cells Which use transcription factors

Promoters are always upstream of the gene for Which the initiate transcription

Promoters can face either direction but synthesis of the new strand is always in the 5' to 3' direction

Elongation

5’ to 3'

RNA polymerase core enzyme DNA add new nucles tides to the 3' end of the growing chain

Termination

specific site called a terminator

DNA encodes this region is such a way that when transcribed the resulting RNA forms a hairpin loop causing the RNA polymerase to stall and fall off the DNA template

Translation

mRNA is a temporary copy of genetic information

Translation decodes the information found in mRNA

  • Protien(polypeptide)

uses ribosomes with accessory protons mRNA and tRNA

Polycistronic messages, translation can result in a group of polypeptides from one message

multiple ribosomes can translate a Single mRNA at one time

  • polyribosomes / polysome

genetic code to decode nucleotide sequences into amino acid sequences.

  • Almost Universal

  • Degenerate - more than one codon for each amino acid

codon - 3 nucleotide Sequence with corresponding amino acid

Decoded using a tRNA which has a complementary 3 nucleotide sequence (anticodon) on one end and an amino acid on the other

nucleotide sequence defines the coding region

Designates the beginning, and end of the region to be translated

70s Ribosomes

RNA and proteins

coordinates peptidyl transfer reaction

Targets for antimicrobial drugs

tRNA

Anticodon recognizes codon and delivers an amins aid

Reversible Changes in tRNA and ribosome for peptidyl transfer

Translation occurs before transcription is complete

  • prokaryotes

Initiation

30s subunit binds to the ribosome binding site

scans to the first AUG to start translation

Initiation complex assembles

  • 30s subunit, fmet tRNA, protein initiation factors

    • Initiating Arna cupies the p-site

50s comes on and initiation factors leave

elongation

A-Site, P-site, E-site

tRNA in the A-site, peptidyl transfer, the ribosome moves forward one codon,

  • tRNA out from E-site

repeat

Termination

Stop codon, does not encode for an amino acid

release factors bind break the covalent bond holding the polypeptide to tRNA and remove ribosomes from message

Prokaryotic vs Eukaryotic

Eukaryotic

  • Transcription and translation ititiation differs

  • Transcription and Translation can't take place at the same time in Eukaryotes

  • mRNA synthesized in a precursor form

    • Pre-MRVA

      • must be processed during and after transcription

  • 5’ end is capped with methylated guanine derivative

    • mRNA stability, proton interaction, translation enhancement

  • 3'end modified by polyadenylation

    • approximate adenine derivatives added to 3’ end of MNA

    • Poly A Tail

    • stabilizes MRNA transcript enhances translation

  • Eukaryotic mRNA contains introns

    • non-coding Sequences

  • Eukaryotic mRNA made in the nucleus

  • monocistronic

  • ribosomes are 80s

Regulation of Bacterial Gene Expression

multiple ways to control gene expression

regulation allows cans to be energy efficient proton regulation can involve inhibition of enzymes or enzyme production usually through transcription

  • reversible

Enzymes can be described by the regulation of their synthesis

Constitutive Enzymes

Always Synthesized.

series always active

enzymes for essential metabolic pathways

Inducible Enzymes

synthesis turned on under certain situations

usually triggered when a certain compound is present such as an energy source

Repressible enzymes

usually synthesized

Genes can be turned off in certain situations

If amino acids become available Synthesis of enzymes in its anabolic pathway is halted

Regulating Transcription

can control one gene, a few genes, or hundreds of genes (global control) at the same time

Alternative Sigma factors

RNA polymerase has sigma factors that fall off after initiation

Alternative Signa factors can recognize different promoters for genes involved in related functions such as response to stress, entering the Stationary phase, or flagella synthesis

Sigma factors themselves can be inhibited by anti-sigma factors

DNA-Binding Proteins

Regions on DNA near the promoter where regulatory protiens can bind to activate or repress transcription

operon - A group of genes controlled by a regulatory protein

Repressors

Regulatory proteins that block transcription

Bind to DNA region-operator-just downstream of promoter and RNA polymerase cannot pass

Allosteric proteins - bind with inducers or corepressors

negative control

  • Induction

  • repression

Induction

Transcription is blocked until an inducer binds the repressor changing the shape of the repressor, and causing it to fall off of the operator

Transcription occurs

Repression

Transcription is on

To turn off repressor

  • the repressor needs to bind the operator But the repressor can not bind alone. A corepressor binds the repressor, changing the shape of the repressor

Binds to operator

Transcription is blocked

Activators

Regulatory proteins that facilitate transcription

Turns it on / allows it to proceed

binding site upstream of a promoter that does not work well on its own

Allosteric proteins - bind inducers

positive control

Lac Operon

controls 3 genes responsible for the breakdown of lactose as an energy source

Both activators and repressors are used so it is a good example of bacterial regulation of gene regulation

Transcription of the operon only occurs if lactose is present and glucose is not

Lactose Converted to allolactose in the cell

The presence of glucose in the cell is monitored by the presence of CAMP

  • inverse relationship

cAMP binds CAP (catabolite activator protein)