Phosphodiester bonds: form between 3’ OH group and 5’ triphosphate, link nucleotides
DNA structure
Anti-parallel double helix
One strand 5’ to 3’, other strand 3’ to 5’
Sugar-phosphate backbone
Base pairs in the inside, held together with H bonds
DNA replication
DNA primer required
Helix unwound by helicase
Replication fork with leading and lagging strand
Leading synthesized in 5’→ 3’ direction - catalyzed by DNA polymerase
Lagging is synthesized in Okazaki fragments which are then joined by DNA ligase
Origins of replication
In eukaryotes, replication starts simultaneously at several points in the genome
Speeds up replication
Bidirectional
Types of RNA
rRNA: combines with proteins to form ribosomes
tRNA: carries amino acids to be incorporated into protein
mRNA: carries genetic information for protein synthesis
RNA polymerases
Multi-subunit complexes which make RNA
Eukaryotes have 3 types: Pol I, Pol II, Pol III
Pol II synthesizes all mRNA
Transcription
TATA box at (-25) - TATA box binding protein introduces a kink into DNA to determine the start and direction of transcription and provides landing platforms for further transcription factors and RNA pol
TFIID - first general transcription factor to bind to the promotor, binds to TATA box through TBP
General transcription factor required for all Pol II transcribed genes
RNA poly II binds specific promotor (0)
Transcription belongs at nucleotide +1
DNA chain separation - unwinding of DNA, catalyzed by helicase
Initiation - selection of first nucleotide of growing RNA
Requires additional general transcription factors
Elongation - addition of further nucleotides to RNA chain in the 5’→ 3’ direction
Termination - release of finished mRNA
Premature → mature mRNA
Splice out introns (exons = coding, intron = non-coding)
Add poly-adenosine tail
Add 5’ cap
Translation
Initiation - formation of initiation complex, energy provided by GTP
Elongation - anticodons of tRNA form base pairs with codons on mRNA, aminoacyl-tRNA synthetases catalyse the covalent attachment of amino acids to their corresponding tRNA molecules
Peptide bond formation and translocation - peptidyl transferase catalyzes peptide bond formation between amino acids in P and A sites, EF-2 moves ribosome along mRNA
Termination - A site encounters stop codon, termination protein binds to the codon and the ribosome dissociates, leads to a change in peptidyl transferase activity which results in the release of the protein from the last tRNA to which it was attached
Post-translational modifications
Glycosylation
Disulphide bods (ER)
Folding/assembly of multi-subunit proteins (ER)
Specific proteolytic cleavage (ER, Golgi, secretory vesicles)
Ribosomes
3 tRNA binding sires - P, E, A
P (peptidyl site) → A (acceptor site) → E (exit site)
Free ribosomes in cytosol produced proteins for cytosol, nucleus and mitochondria
Post-translational - produced in cytosol then translocated (after translation)
Bound ribosomes on rough ER produce proteins for plasma membrane, ER, Golgi, secretion
Co-translational - translocation concurrent with translation
Mutations
Point mutations: change in single base in DNA
Missense mutation: results in change of amino acid sequence
Nonsense mutation: creates new termination codon
Silent mutation: no change of amino acid sequence
Frameshift mutation: addition or deletion of 1 or 2 bases which changes the reading frame of translation
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