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51 Cards in this Set

  • Front
  • Back
Nucleotides

Composed of what? And what do they form?
Composed of:
1. Nitrogen base
2. Ribose Sugar
3. Phosphate Group

Form a polymer of nucleotides or nucleac acid (DNA or RNA)
Types of Nucleotides?

What are the base pairs?
Types
1. Purine
a. Adenine
b. Guanine

2. Pyrimidine
a. Cyostine
b. Thymine
c. Uracil (RNA Only)

Base Pairs
1. A and T = 2 H Bonds
2. G and C = 3 H Bonds
DNA Structure of Prokaryotes (Bacteria)

Supercoiling?
1. DNA structure is circular, and has the ability to pack in the DNA through Supercoiling.
DNA Structure of Prokaryotes

Supercoiling - How does it occur?
Through topoisomerases (enzyme)

Topoismerases create a break in the photodiester bond of a single or double strand allowing for a coiling.

1. Topoisomerases I = Makes single stranded breaks

2. Topoismerases II (DNA Gyrase) makes double stranded breaks
DNA Structure of Prokaryotes (Bacteria)

Stemloop Structure
1. created by inverted repeats. Cruciform structure forms from hydrogen bonding between a single DNA strand.

2. Structure serves a protein-binding site.
Different Forms of DNA

Eukaryotes?
1. Linear

2. Wrapped around histones (forms nucleosomes)

3. Found in the nucleus

4. Diploid = 2 copies of the chromosome

5. Telomeres – no coding sequences that repeat
Different Forms of DNA

Prokaryotes
1. Circular molecule

2. Packaging lacks histones

3. Found in cytoplasm

4. Nucleoid

5. Hemizygous – single copy of a chromosome
Different Forms of DNA

Eukaryotes

Histones
Small positively charged proteins that neutralize the negative charge of DNA.

Used to help coil and compact the linear DNA of eukaryotes and some Archae
Replication

Prokaryotes (Bacteria)

Steps
Growth proceeds from 5'-phosphate to the 3'-hydroxl end.

Steps
1. DNA is unwound by the enzyme helicase, separating. This happens on two ends, creating a replication fork on both ends.

2. Single stranded binding proteins latch on to keep seperate.

3. Primase synthesizes a short RNA segment called the RNA Primer

4. a. Leading Strand - DNA Polymerase attaches onto the end of the Primer '3 -OH group and starts to catalyzes '5-phosphate to the hydroxyl end and continues on.

4. b. Lagging Strand - Similar to leading strand, but because there is no 3-OH readily available the Primase has to continully make a primer that the DNA polymerase III can link to.

5. Lagging Strand - DNA Poly II continues until it reaches synthesized DNA, then it stops

6. DNA Poly I then takes over and continues synthesizing while removing the Primer

7. Then DNA Ligase comes in and seals up the nicks.
Replication

Eukaryotes

Replication of Linear DNA Requires what?
a DNA Primer with a -OH group attached is used to initiate synthesis from the DNA polymerase III.
Trasncription

What is it? What is the difference from DNA?
1. Synthesis of RNA

2.
a. RNA contains ribose, not deoxyribose
b. RNA contains uracil
c. RNA is not double stranded
Transcription

Bacteria

3 Broad Categories
1. Initiation
2. Elongation
3. Termination
Transcription

Bacteria

Steps of Transcription
1. RNA Poly reads DNA until it finds a region for which it has high affinity, called the Promoter region

2. RNA Poly recognizes a promoter with assistance from accessory proteins, the most significant is the Sigma Factor.

3. RNA Poly unwinds a portion of the DNA double helix

4. Initiation - RNA Poly synthesizes a dinculeotide and then begins to travel and synthesizes the complimentary RNA molecule.

5. Once a short strand is synthesized the Sigma Factor falls away.

6. Elongation - RNA Poly synthesize nucleotide monomers into an RNA polymer using protenis called Elongation Factors
- RNA Poly re-zips the DNA

7. RNA Poly encounters a second type of consensus sequence telling it to stop polymerizing or
Transcription

Bacteria

Sigma Factors
Proteins that identify certain promotor regions, specifically -35 sequence and Pribnow Box (-10 Sequence), that are just upstream from the start of the transcription site.
Transcription

Bacteria

Ways from Termination (2 Ways)
1. DNA inverted repeat that causes the synthesized RNA to make a stem loop. The RNA Poly stops at the Stem Loop and falls off, and the DNA and RNA come apart.

2. Rho Dependent Termination
a. Rho binds to RNA
b. Moves along molecule until it reaches RNA POL
c. RNA Poly pauses at Rho dependent termination site
d. Rho causes RNA POL to fall off DNA, ending transcription.
Transcription

Bacteria

Transcription produces 3 types of RNA
1. Messenger RNA
2. Transfer RNA
3. Ribosomal RNA
Transcription

Messenger RNA (What is its purpose?)
-Contains an RNA version of the DNA found within a gene

-The information in mRNA is directly translated into a protein.

- In Eukaryotes, mRNA goes through a maturation process before it is function
a. Introns (non-coding regions) are removed and exons (coding regeions are combined) by the Splicsome (Protien and snRNA)
b. Also a cap and tail are put on

* Prokaryotes do not have as many nitrons, and they don't cap or have a poly A tail.
Transscription

Eukaryotes

mRNA

5' Cap
The 5’ end of Eukaryotic RNA contains a modified Guanosine residue which delays degradation of transcription and promotes the formation of the initiation complex between the mRNA and ribosome.
Transscription

Eukaryotes

mRNA

Poly A Tail
1. The 3’ end of Eukaryotic mRNA contains 100-200 residues of the nucleotide Adenine.

2. Enhances export of mRNA from the nucleus

3. Prevents degradation in cytoplasm

4. May enhance translation
Transcription

Transfer RNA
1. tRNA lacks a genetic message but instead acts as a structural element involved in the translation of mRNA sequences.

2. Short molecule (73-93 nucleotides long)

3. Contains 4 stems and 3 loops

4. Interacts both with Amino Acids and mRNA
Transcription

Transfer RNA

Codon, Start Codon, and Stop Codon
sequence of 3 nucleotides in an mRNA molecule which calls for a specific amino acid to be incorporated into a growing protein..

Start Codon - Usually AUG that signals the start of a protein

Stop - Codon that signals the end of a protein
Transcription

Transfer RNA

Redundancy and Codon Bias
more than one triplet may code for the same amino acid. Example- ACU, ACC, ACA all encode for Threonine. All codons for a particular amino acid may not be used equally; one codon may be the preferred sequence.

Codon Bias - one organism may prefer one codon over another to make an amino acid.
Transcription

Transfer RNA

Recognition, Activation, and Charging of tRNAs

Steps
1. Activation of the amino acid by reaction with ATP crreates an aminoacyl-AMP, which remains attached the synthase.

2. Then the activated amino acid is attached to the tRNA to create a charged tRNA

3. Then the charged tRNA goes to the ribosome to charge the tRNA
Transcription

Transfer RNA

Recognition, Activation, and Charging of tRNAs

2 Points
1. Specific Amino acyl tRNA synthase for each codon AA/combination

2. Enzyme recognizes the anticodon region of the tRNA and the amino acid
Transcription

Archaea
1. Similar in bacteria in that RNA polymerase is used, but it is intiated by three promoters:
a. BRE
b. TATA
C. INT

2. Te TATA-binding protein binds to the TATA promoter and the Transcription Factor B binds to the INT and BRE promoters.

3. Once in place the RNA Poly will bind and start transcription
Transcription

Eukaryotes

Difference from Archea and Bacteria
Eukaryotes have three RNA polymerases that transcribe the major subclasses of RNA

1. RNA Polymerase I- most types of rRNA

2. RNA Polymerase II- all mRNA

3. RNA Polymerase III- tRNA (and 1 rRNA)
Transcription

Eukaryotes

RNA Polymerase II Transcription
The RNA Poly II binds directltly to the INT and the TATA-binding protein binds to the TATA promoter
Translation

Ribosomes
1. Complex of proteins and rRNA (Ribosomal RNA)

2. Consists of 2 subunits- Small subunit and Large subunit
Translation

rRNA

Purpose
1. Purpose
a. Contains no genetic message
b. Acts as a structural and catalytic element in the translation of mRNA into protein.
Translation

tRNA

Types and What They Do.
1. 16s rRNA-
a. helps 30s sub unit identify Shine Delgarno sequence of mRNA.
b. physically attaches tRNA via anti-codon loop to ribosome.
c. participates in recognition of Release Factor proteins.

2. 23s rRNA
a. recognized acceptor stem of charged tRNA
b. catalyzes formation of peptide bonds
c. aids in translocation
Translation

Four Steps
1. Initiation
2. Elongation
3. Translocation
4. Termination
Translation

RIbosomes
Property Prokaryote Eukaryote
Overall Size 70s 80s
Small Subunit 30s 40s
# Proteins Small su ~21 ~30
RNA Size 16s 18s
Large Subunit 50s 60s
# Proteins Large su ~34 ~50
RNA Size 23s, 5s 28s, 5.8s, 5s
Translation

Initiation

Prokaryotes

Steps
1. 30 s (small subunit) binds to docking site (Shine Delgarno Sequence (3 to 9 nucleotides)) present in mRNA with help of Initiation Factors. 16s rRNA hybridizes to SD sequence.

2. tRNA-met binds to 30s s.u. and mRNA (This step requires energy in the form of high energy phosphate bonds contained within GTP)

3. Initiation Factors (proteins) recruit 50s (large) sub unit. At this point the intact ribosome has been formed with tRNAmet in P site.
Translation

Initiation

Prokaryotes and Eukaryotes

tRNAmet What does it do?
1. tRNAmet is the first AA in eukaryotic protein synthesis (formyl-methione in prokaryotes)

Formyl group or entire amino acid may later be removed in post-translational modifications.
Translation

Initiation

Intact ribosomes contain how many sites? Name?
1. A site (acceptor site)
2. P site (peptide site)
3. E site (exit site)

*tRNAmet (initiation codon) found in what is referred to as the P site (peptide site).
Translation

Initiation

Start Codon
1. Usually AUG

2. GUG may sometimes serve as an alternative initiation codon
Translation

Elongation

Steps
1. The tRNAaa hybridizes to mRNA at A Site via hydrogen bonding

2. RNA catalyzed condensation reaction creates dipeptide attached to tRNAgly-met.

3. Now an uncharge tRNA is still on the P site, while a tRNA with a peptide chain is on the A site

* Remember, this requires GTP and elongation factors
Translation

Translocation
1. The dipeptide tRNAgly-met now occupies the P site of the ribosome.

2. A site is now vacant, ready for the next tRNA molecule specified by the codon in the mRNA.

3. During subsequent translocation uncharged tRNA once linked to methionine discharged from the ribosome.
Translation

Translocation

Getting Stalled
If this occurs the ribosoms has the TmRNA, which will cleave the UUU tRNA and have its own UAG stop codon that will initiate disassembly of the protein structure
Translation

Translocation

Termination
1. Release Factors enters A site and binds to nonsense or stop codon

2. Release Factors hydrolyzes tRNA-protein bond, releasing protein

3. Ribosome dissociates into subunits + mRNA
Translation

Translocation

Polysomes
Multiple ribosomes translating same mRNA molecule
Proper Folding
1. All proteins must be folded in the proper confirmation to be active.

2. Chaperonins (ATP dependent protein) may assist in this process.
Proper Folding

Chaperonins

Types
1. DNA J and DNA K - They function in the cell to prevent a protein from folding too quickly and possibly improperly.

2. GroEL and GroES - These proteins may help fold proteins that DNA J and DNA K were unable to fold. They also may help re-fold proteins that are partially denatured.
Protein Secretion

signal sequence
may signify the secretion of a protein or the insertion of that protein into the membrane.
Protein Secretion

Unfolded

Signal Recognistion Particle (SRP) and SecA
1. SRP - compose of protein and RNA binds the signal sequence and facilitates entry of the growing peptide into the membrane bound transport protein.

2. SecA - Binds through the membrane

* BOTH require ATP
Protein Secretion

Folded

TaT
1. moves a protein across the membrane after it has been properly folded. Tat protein export uses the Proton

**Proton Motive force to provide the energy required for export.
Protein Splicing
1. has been observed, analogous to mRNA splicing in Eukaryotes where certain regions are removed (Inreins) and other regions are covalently linked together (Exteins) to form the mature protein.

2. DNA Gyrase subunit A in Mycopbacterium leprae undergoes such processing.
Control of Gene Expression

Occurs at what levels?
1. Initiation of transcription
2. Elongation of transcription
3. Initiation of translation
4. Post translational modification
Control of Gene Expression

DNA Binding Proteins

What are they?
1. Proteins that interact with DNA often have characteristic motifs that allow the protein to recognize specific sequences in the DNA.

2. Most DNA binding proteins read the Major Groove while some read the Minor Groove of the molecule.
Control of Gene Expression

DNA Binding Proteins

Structure

Helix Turn Helix
Motif contains a stabilizing helix and a recognition helix.
Control of Gene Expression

DNA Binding Proteins

Helix Turn Helix

Leucine Zipper
The two stabilizing helices interact via hydrophobic interactions between leucine residues spaced every 7 amino acids.

* Helps to hold the helicases together