Genetics 1

• ATP is made up of adenine and ribose, which make adenosine, and three phosphate groups
• DNA = Deoxyribonucleic acid
• RNA = Ribonucleic acid
• The difference in structure of DNA and RNA:

  • DNA has one OH group on 3 prime carbon but when in a chain this OH group isn't present
  • RNA has 2 OH groups one at 2 prime and one at 3 prime end and when in a chain there is still an OH group
  • DNA is more stable than RNA
• Within chains of DNA and RNA there is an alternating sugar phosphate backbone with a base pointing up. The neighbouring nucleotides are joined together via phosphodiester bonds.
• The phosphate group on nucleotides is what gives DNA and RNA a negative charge
• Purines - Adenine and Guanine
• Pyrimidines - thymine , cytosine and uracil
• Uracil differs from thymine by the attachment of hydrogen instead of a methyl group to the 5 prime carbon in pyrimidine ring.
• Bases A,T,C,G and U are attached by β glycosidic bonds. C-1' of sugar attached to the N-9 of the purine or the N-1 of pyrimidine.
• Nucleoside triphosphates are the building blocks of DNA & RNA
• Phosphoryl group attached to the 5 prime carbon atom of the sugar and free hydroxyl group is on 3 prime end of the carbon. Code is always written from 5' to 3' direction.
• Adenine and Thymine form a base pair via two hydrogen bonds, guanine and cytosine form a base pair via three hydrogen bonds.
• DNA has:

  • antiparallel polynucleotide strands
  • 1 : 1 of pyrimidine : purine
  • right handed helix with bases inside
  • major ( sides of bases are exposed ) and minor ( little exposure ) groove
• Before RNA is produced, there are promoter sites along the DNA template strand with specific bases. One promoter site is found at -25 position and is called the TATA box and another less frequent site is found at -75 position and is called the CAAT box. These promoter regions act as binding sites for proteins that start the production of RNA.
• Genes contain exons (protein encoding) and introns (connecting)
• Primary transcripts contain introns & exons and are modified with a cap at 5' and at the 3' end adenine residues to stabilise the RNA in the cell.
• Introns are spliced out from primary transcript by spliceosomes to generate mature messsenger RNA.
• Base pairs can be recognised by their side groups
• Proteins can interact with single stranded DNA and prevent base pairs forming within a strand and creating ' hairpins ' that stop the action of DNA polymerase.
• Eukaryotes - Cytosine methylation:

  • On DNA
  • addition of methyl group
  • at CG motifs
  • Maintanence methyltransferase
  • if parent cell is methylated daughter cell will also be methylated at CG site and vice versa
  • CpG islands ( p is for phosphate) important in regulating gene expression
• Cytosine methylation allows cell to identify original template strand from new strand, as only the original is methylated
• In prokaryotes there are 3 different types of methylation:

  • N4-methylcytosine (4mC)
  • 5-methylcytosine (5mC)
  • N6-methyladenine (6mA)

  This regulates gene expression and virulence of attacking bacteria and pathogen-host interactions, making them more invasive
• In prokaryotes methylation is important for DNA replication:

  E. coli DNA replication

  • origin of replication that is 250 base pairs
  • contains N6-methyladenine in 11 GATC sequences within 250 bp
• E. coli DNA replication :

  • G1 phase (full methylation) - at 11 sites and further down structure will have methyl group that stimulates expression of protein from gene dna A
  • G1/S phase - dna a will bind to origin of replication at methyl sites and start dna synthesis and break apart strands
  • Early S phase - generation of 2 strands one methylated the other not, overall hemi-methylated. SecA binds to h-m sites and also to end site to turn of dna A to end DNA replication
  • S phase re-methylation - DAM protein methylates new strand and returns of origin of replication back to original state
• DpnI in the lab specifically cleaves double stranded dna where it is methylated on both sides
• Wild type DNA sequence can have DNA damage in form of base loss, alteration , mismatch or crosslink and strand breaks.

  This can be caused by errors in DNA replication and Mutagens
• DNA damage can be recognised by body cells and allow for repair by direct reversal and multi-step repair pathways returning it to wild type DNA sequences.
• If DNA damage isn't detected or repair is not successful, the result is a mutant dna sequence. Mutations can be beneficial, harmful or neutral.
• Mutations:

  • A -> G and C -> T and vice versa are called transitions
  • A -> G and G -> C and vice versa are called transversions
• DNA mutation damage types:

  • Deamination -> removing nitrogen groups
  • Oxidative damage -> due to reactive oxygen group
  • Depurination -> cleaving off purine
  • Alkylating agents -> bind to nucleotides
  • Bulky adducts -> bind to nucleotides
  • Base analogues -> resemble nucleotide bases
  • Intercalating agents -> incbetween bases prevent dna replication
  • UV light -> can cause crosslinking between bases, lead to blocked replication
  • Ionizing radiation -> gamma and x-rays that break sugar phosphate backbones
• Ribonucleic acid (RNA)

  • ribose sugar
  • phosphate group
  • Nitrogenous base (A,U,G or C)
  • form secondary structures
• RNA's secondary structure relates to its function:

  1. Ribozyme - > allows specific site cleavage
  2. Ribosomal RNA -> ribosomes are made largely of RNA (rRNA)
  3. Transfer RNA (tRNA)-> aid formation of proteins by bringing single amino acids to ribosome
• tRNA contains over 50 different types of modified bases usually represented by symbols. These modified bases make up around 10% of tRNA and are modified in the nucleus.
• Modified nucleotides have a few functions including :

  Structural

  • change base pairings
  • hydrophobicity
  • involved in protein interactions
• Wobble base-pairing position is found at 5' end of anticodon on tRNA and 3' end of codon on mRNA. Due to wobble position even if there is a change in the third base in a codon, it will still code for the same amino acid.
• In DNA genomes contain many repetitive sequences and can form structural binding sites.
• Function of DNA repeats/Secondary structures:

  • replication
  • recombination
  • repair
  • transcription
• DNA origami can be used to make molecular cages that could possibly contain drugs that can be targeted to a particular cell or part of the body.
• DNA denaturation -> double stranded base pairs can be separated by temperatures of 100 degrees or a high pH (over 13)