biochem - unit 2 - DNA & RNA

Adenine. A nitrogenous base. Is a purine (2 rings) and has only an amino group.

Guanine. A nitrogenous base. Is a purine (2 rings). Has an amino group and a carbonyl

Cytosine. A nitrogenous base. Is a pyrimidine (only one ring). Has an amino group and a carbonyl

Thymine. A nitrogenous base. Is a pyrimidine (only one ring). Has two carbonyls and a methyl group

Uracil is a nitrogenous base. Is a pyrimidine (only one ring). Has two carbonyls.

Nucleoside

This is the coupling of a nitrogenous base with a sugar

Nucleotides

This is the coupling of a nitrogenous base with a sugar and a phosphate(s)

What are the properties of DNA double helix-7

Two complementary strands, antiparallel, sugar-phosphate backbones (polar) face outward towards the water, bases sit in the inside of the structure, hydrogen bonding (A-T, have 2 bonds) & (C-G, HAVE 3 BONDS), base stacking (van de waals interaction), hydrophobic effect (the bases are nonpolar and want to face the inside, and creates a stabilizing effect), major and minor groups. B DNA form, 10- 10.5 bp per turn, and 2 nm in diameter.

Complementary strand sequence

A←→T and G←→C. The strands are antiparallel. The BP of the complementary strand needs to match the given strand in the 3’—>5’ direction, because the given strand will run in the 5’—>3’ direction, because of the way it is aligned beneath it

differences between DNA & RNA

Sugar—> DNA: deoxyribose(H instead of OH @ C2’) & RNA: ribose( OH @ C2)

Bases—> DNA: A, T, C, G & RNA: A, U, C, G

Structure—> DNA: normally double-stranded & RNA: usually single-stranded, and can fold into paired regions, hairpin, cloverleaf, and hairpin

main role—> DNA: stores genetic information & RNA: regulates gene expression: mRNA, tRNA, rRNA

Stability—> DNA: more chemically stable because there is no oxygen @ C2’ & RNA: less chemically stable bc it is more susceptible to hydrolysis because it has a hydroxyl group @ C2’

Restriction Endonuclease

An enzyme that cuts DNA @ particular recognition sites in DNA. Restriction endonucleases recognize palindromic DNA sites (read the same backward as it does forward), this is where a DNA region is cut.

Polymerase Chain Reaction - PCR

A method that is used to copy regions of DNA. It amplifies a chosen DNA region. It needs a template of DNA, a thermostable DNA polymerase, 4 dNTPs, and a suitable buffer/ions. 2 primers will go and mark the boundaries of the DNA segment that needs to be amplified. Both primers will point their 3’ ends into the region.

1. Denaturation—> heat will separate the DNA strands.

2. Annealing—> cooling will allow primers to form base pairs with their targets and create a border

3. Extension—> DNA polymerase can extend in the 5’—>3’ direction and make new DNA.

Repeated cycles will ideally double the amount of the targeted DNA, roughly 2ⁿ amplification after n amount of cycles. Realistically, the reaction will become less efficient over time and plateau. Primers and dNTPs become limited, polymerase loose their activity, and newly amplified DNA will pair with itself instead of the primers

Replication

DNA—>DNA. This process makes copies of DNA before cell division. Happens in the nucleus

Transcription

DNA—>RNA. RNA polymerase will use DNA as a template to make an RNA copy of a gene

Translation

mRNA—>protein. Ribosomes will use codons to build an amino acid chain that is later folded into an active protein

Semiconservative replication

The parental strands will separate and each will become a template for its new partner. This means that each daughter strand will contain one parental strand and one newly synthesized strand

5’—>3’ synthesis

Although proofreading can be conducted in a 3’—>5’ direction. Synthesis is conducted in a 5’—>3’ direction. Nucleotides are added to the nucleotide’s 3’ end, and the newly added nucleotide provides a new 3’ OH

DNA replication: template primer

This is the starting setup that DNA polymerase needs to begin replication. The template is the already existing DNA strand that tells DNA polymerase what bases to add. The primer is a short strand of DNA that is attached to the template; it supplies a free 3’ OH

Transcription Mechanism: template starnd

In transcription, RNA polymerase will use one strand of DNA (template strand) to make RNA. This is for the expression of specific genes.

1. DNA will open locally to expose the template, section of the gene

2. RNA polymerase will then read the template in a 3’—>5’ direction

3. RNA polymerase can then add the complementary RNA nucleotides that make RNA in the 5’—>3’ direction

RNA polymerase does not need a primer

mRNA

Messenger RNA: this is the strand that is taken to the ribosomes. It carries the coding information to the ribosomes. has/makes the codons.

tRNA

Transfer RNA: This is the small RNA molecule that will act as the adaptor for the genetic code in mRNA. has the anticodon sequence that is complementary to mRNA. mRNA—tRNA—amino acid sequence

rRNA

Ribosomal RNA. The structural and catalytic part of the ribosomes. It is non-coding. Helps catalyze the formation of peptide bonds in protein synthesis. secondary structure is a cloverleaf and its tertiart structure is -shaped

RNA modification

These are RNA modifications that happen after transcription has taken place. These modifications help it become functional:

• Cutting and trimming- pre-RNA is trimmed and shortened to its proper size

• Chemical modification- some of the bases are changed, like inosine (adenosine) or 5-methylcytidine (a methyl group at the 5’ C of cytosine) in tRNA.

• Splicing- introns are removed from the sequence, and exons are joined in eukaryotic pre-mRNA

• Adds a cap and tail- eukaryotic mRNA will receive a 5’ cap and a 3’ poly(A) tail; this is to help protect these structures and support their use

These changes help process the RNA; they don’t turn it into the protein

tRNA structure

This is a structure that helps convert mRNA—>amino acid sequence.

• A short chain of RNA

• Its secondary structure is a cloverleaf

• Its tertiary structure is L-shaped

• Has modified bases like inosine (adenosine) or 5-methylcytidine (a methyl group at the 5’ C of cytosine) in tRNA.

• Paired stems: complementary bases in the single strand of RNA can hydrogen bond together to form short double-stranded regions

• It has a 5’ phosphorylated end

• 3’ CCA end will attach to the amino acid on the terminal adenosine

• The anticodon region of tRNA will pair with the codons of mRNA

Genetic code

This is the set of rules that are used to map mRNA nucleotides into amino acids:

• Each codon only has 3 bases

• In each strand of RNA, there are 64 codons: 61 are used to specify amino acid sequence, and 3 are used to signal stop

• AUG is used to code for methionine and usually serves as the start codon

• Multiple codons can code for the same amino acid- redundancy or degeneracy

• Codons are only read consecutively, without overlap, and in an established reading frame

• This code is nearly universal across all organisms

a codon is 3 mRNA bases read 5’—>3’

Protein synthesis steps

1. Initiation—> mRNA will bind to a ribosome. A specific initiation mechanism is needed to start protein synthesis. This means that the ribosome has to find and assemble at the right codon/the start codon of an mRNA strand before it can start making proteins

2. Aminoacyl-tRNA will bind one after another to the next codon sequence of the mRNA chain

   • Aminoacyl-tRNA is a tRNA with an amino acid attached.

   • Encounters are random, but acceptance can be checked through codon-anticodon pairing

3. Elongation—> an amino acid is added to the C-terminus of a peptide in a peptide chain. Peptide bond synthesis is catalyzed of formed by rRNA

4. Translocation—> the ribosome moves on to the next codon

5. Termination—> when a stop codon is encountered by tRNA or a release factor; the completed protein chain is released

ribose