DNA/RNA/Proteins: Replication/Transcription/Translation

Bases

Pyrimidine (1 ring)

Cytosine

Thymine

Uracil

“Cut the Py”

Purines (double ring)

Adenine

Guanine

AT(u)—2 H bonds

GC—3 H bonds

Uracil — RNA

GCs, more H bonds needs a lot of heat in PCR (amplification/replication) to break them

Prokaryotes

1. Methylation (restriction enzymes act like an immune system and will chop up DNA and restrict growth of viruses whose DNA is NOT methylated)

2. Super coiling (DNA gyrase)

3. Plasmids (extrachromosomal, five bacteria extra abilities like antibiotic resistance)

DNA charge and what does it wrap around?

DNA is negatively charged and it wraps around histones. Histones contain basic (positive) amino acids to attach to -DNA.

Nucleosome,Chromatin,Chromosomes? Euchromatin vs Heterochromatin

Nucleosome=Histone + DNA

Chromatin=Packaged nucleosome

Chromosomes=2 sister chromatids (replicated)

Euchromatin = unwound, active, light staining (E for Expressed)

Heterochromatin= tightly wound, inactive, dark staining (H for Hibernating)

What are Telomeres?

Protective caps made of repetitive DNA sequences at the end of chromosomes

Central Dogma

DNA-(transcription)-RNA-(translation)-Proteins

Reverse transcription (Retroviruses like HIV)

What are codons?

3 nucleotides=1 amino acid….. “words”

Start? Stop? Codons

Start-AUG Met

Stop: UAA, UGA, UAG

What are intergenic regions?

They are non-coding spaces/stretch of DNA between genes on a chromosome.

Mutation types

1. Polymerase errors (point mutations, small repeats, insertions/deletions)

2. Endogenous damage (reactive oxygen species ROS, physical damage, causes oxidized DNA. crosslinked, breaks)

3. Exogenous damage—external (radiation and chemicals) (UV radiation, X rays)

4. Transposons “jumping genes”

***PPP in metabolism produces NADPH which can help to neutralize ROS

Point mutations—single base pair change

1. Missense: Codon for aa becomes new codon for new aa (change depends on what to what properties)

2. Nonsense: Codon aa becomes stop (STOP THAT NONSENSE)

3. Silent (codon for aa becomes new codon for same aa)

Transposons

transposase enzyme —cut and paste DNA

Contribute to random variation

If inserted into intergenic region no impact vs coding region can disrupt gene or cause mutations

Bad bases, Mis match repair pathway

-during or shortly after replication (narrow window)

-Parent is methylated daughter is not so recognition happens

-Endonuclease comes in and repairs without causing harm

Bad bases, base/nucleotide excision repair

-Can happen at any time so wide window

-Do not know which one is bad so can lead to mutations

Broken chromosomes, homology directed vs non-homologous directed repair

Homology: After replication, use identical sister as a template, repair causes no loss of DNA, only G2 phase of cell cycle

Non-homologous: Any time during cell cycle, mutagenic possibly, and translocations (swapping) can happen, can cause some harm

DNA Replication Rules

1. Semi conservative

2. Made 5’—3’, read 3’—5’

3. Primer is needed

DNA replication enzymes

1. Helicase: unzips and unwinds DNA

2. Topoisomerase: Single stranded, cuts, releases tension, and relaxes super coiling

3. Primase: Lays down RNA primer

4. DNA polymerase: Replicates DNA, proof reads

5. Ligase: links okazaki fragments

Leading strand is what is made normally

Lagging strand: DNA polymerase flips it , and overlap forms, chunks on lagging strand (okazaki fragments linked by ligase)

Prokaryotic DNA replication

5 DNA polymerases

I and III are for replication

II,IV, V are for repair

single orgin

Eukaryotic replication

Multiple origins

Replication bubbles

Several DNA polys, multisubunit enzymes

Telomeres

With each round of DNA replication, DNA gets slightly shorter (AGING)

Telomerase elongates telomers on parent strands on DNA

Cells that express telomerase “immortal” high turnover like stem cells and cancer cells

DNA VS. RNA

DNA: Double stranded, thymine used, deoxyribose, double helix

RNA: Single, uracil used, ribose, lots of shapes

Types of RNA

rRNA, mRNA, and tRNA

miRNA/siRNA: Can inhibit translation, and mark RNA for breakdown

Transcription

DNA—RNA

5’—3’ made

No primer needed

Template is non coding strand and this is what is read, mRNA is complimentary to non-coding strand

***RNA polymerase makes mRNA from template

Prokaryotes vs eukaryotes transcription and translation

Prokaryotes: These happen at SAME time, no mRNA processing, polycistronic meaning many different proteins from single mRNA, 1 RNA polymerase for transcription

Transcription: Yes, both eukaryotic and prokaryotic RNA polymerases require a DNA template to synthesize RNA. They both follow the same fundamental mechanism, reading the template DNA strand in the direction to produce a complementary RNA molecule. The primary difference lies in how they locate the template: prokaryotes use a single RNA polymerase with a sigma factor, whereas eukaryotes require multiple, distinct RNA polymerases and general transcription factors to initiate the process.

Eukaryotic: Diff places and times, mRNA processing (Gcap 5’, poly A tail 3’, splicing (EXons are expresses and INtrons go in the trash), 1 mrna=1 proteins, RNA polyemerases

RNA polymerase I-rRNA

II-mRNA

III-tRNA

Euk RNA polymerasese are empty

are=r I

M=II

t=III

tRNA and mRNA

anticodon (tRNA) for mRNA

Aminoacyl tRNA synthase: attaches amino acid to tRNA

Wobble base pairing: : Occurs at the 5' base of the tRNA anticodon. This phenomenon allows a single tRNA to recognize multiple codons by "wobbling" at the third position.

Ribsomes

Prok: subunits large=50s small=30s total 70s

Euk: subunits large=60s small=40s total 80s

Prok are odd, 3,5,7 are odd

Euk are even, 4,6,8 even

+20 between the number and 10 between the two types prok smaller

Translation

E site: exit site for empty tRNA

P site: growing Protein held here

A site: new Amino acid here

Initiation: Small subunit binds the mRNA met-tRNA attaches to P site, large then attaches

Elongation: AA-tRNA binds at A site. Ribosome catalyzes formation of peptide bond and transfers the protein to AA in A site.

Ribsome moves forward one codon and puts the empty tRNA in the E site, the protein is now in the P site attached to the tRNA. The A site now open for next amino acid tRNA. Keep doing this until stop codon in A site ,

Translation is the biological process where ribosomes in the cytoplasm or rough ER read messenger RNA (mRNA) sequences to build protein chains. It converts genetic code, in three-base "codons," into a sequence of amino acids, occurring in three stages: initiation, elongation, and termination.

Overall ATP for translation

number of amino acids * 4= ATP needed

tRNA loading requires the most ATP

Post Translational Modifications

Protein folding

—chaperones help with this

covalent mods

—disulfide, glycosylation, phosphorylation

processing

—cleavage to form active protein