OECG

TRANSCRIPTION IN EUKARYOTES General process: Initiation, elongation and termination Initiation Enzyme involved: RNA polymerase II proteins involved: General Transcription factors ● TBP ( Tata binding protein) ● Other general transcription factors So what does TBP do? ● TBP binds to promoter at the TATA box ● Recruits other GTF which helps with binding of RNA polymerase to bind to the promoter ● Forming preinitiation complex What does RNA polymerase do? ● Unwinds and unzips the DNA strands Why does RNA polymerase need to do that? ● To expose the nitrogenous base so that it can act as a template for transcription ● For complementary base pairing for free nucleotides Elongation Enzyme involved: rna polymerase What does rna polymerase do? ● Reads the DNA template from 3’ to 5’ direction ○ The rna synthesised from 5’ to 3’ ○ Free nucleotides are paired with their complementary bases via hydrogen bonds ● Catalyses formation of phosphodiester bonds between a 3’ hydroxyl end to incoming 5’ phosphate group Termination Enzyme involved: RNA polymerase Products formed: Pre-mRna

Whats the terminator sequence called: polyadenylation sequence signal What does RNA polymerase do? ● transcribes a polyadenylation code AAUAAA past its signal sequence ● the growing chain is then Cleaved downstream 10-30 nts, RNA dissociates from DNA What happens after transcription is done? ● RNA transcript is released ● RNA polymerase dissociates from DNA Pre-mRNA modification (only in eukaryotes) Why is it important? ● to produce mature mRna ● helps with translation Note: can occur simultaneously with transcription Addition of 5’ caps (ALWAYS THE FIRST) when does it occur? ● during synthesis of pre-mRNA Enzymes involved: capping enzymes What does it do? ● Capping enzymes catalyse the covalent attachment of 7-methylguanosine to the 5’ end of the mRNA transcript, forming a 5’-5’ triphosphate linkage. Why does it do it? ● protects mRNA from degradation by 5’ exonucleases, conferring stability to the capped mRNA ● serves as a binding site for translation initiation factors and ribosome to initiate translation. ● facilitates the export of mature mRNA from the nucleus to the cytoplasm.

addition of 3’ poly(A) tails process: Polyadenylation Enzyme involved: poly-(A)-polymerase What does the enzyme do? ● The enzyme catalyses the addition of 50-240 adenine nuecleotides to the 3’ end Why does the enzyme do that? ● to facilitate the transport of mRNA from nucleus to cytoplasm ● prevent degradation by the 3’ exonucleases ● facilitate binding of ribosomes to 5’ end of mRNA and initiate translation Splicing PROTEINS involved: splicesome What does this protein do? ● splice out the introns from the mRNA strand How does the protein do that? ● recognises the 5’ splice site, branch point, 3’ splice set ● excise the introns and ligate the exons Why does it do that? ● introns allow alternative splicing to occur so that multiple different mRNA can be produced from a single gene ● to prevent addition of nucleic acids not needed which can affect the protein structure due to unfavourable R group interactions TRANSLATION IN EUKARYOTES general process: initiation, elongation, termination Things to note ● Sequences of bases in mRNA provides coded information that is read by ribosomes in 3 consecutive nucleotides known as codons

● Characteristic of codons ○ Codons are universal as they are common in most organisms ○ Punctuated: consists of codons that signal when the sequence stops and begins ■ Start codon: AUG ■ Stop codon: UGA, UAG, UAA ○ Codons are non-overlapping as the same nucleotide are not used for more than 1 codon ○ Unambiguous as no codons code for more than one amino acid ○ Degenerate as One amino acid can be produced by more than one codons as codons have more than one tRNA and some tRNA can tolerate mismatch at the 3rd base of a codons Initiation Enzymes involved: What does the enzyme do? Plasmid are not part of the chromosomes TRANSCRIPTION IN PROKARYOTES TRANSLATION IN PROKARYOTES

ORGANISATION AND CONTROL IN EUKARYOTES Differential expression: the process where cells become different from one another due to the unique sequences of genes that are expressed Why is this important? ● So that they can produce different cell types in a single organism to carry out their specific functions from the same genome ○ Eg) liver cells, nerve cells ● Enable multicellular organisms to proceed through development stages ○ As organism progress through developmental stages, certain genes are activated or repressed at certain stages but not others. Why do we need to regulate transcription and translation? ● To save energy and resources especially when the gene is not needed DNA Packing 1st stage: Proteins involved: histone proteins and Linker DNA Products formed: 10nm nucleosome Structure formed: beads-on-the-string What do histones protein do? ● DNA winds around histone octamer containing 8 histones proteins ● Consist of high proportion of positively-charged amino acids, allowing them to bind tightly to the negatively-charged phosphate groups along DNA backbone via ionic and hydrogen bonds What does linker DNA do? ● Connect nucleosome together giving rise to bead on string structure, shortening the length of DNA

2nd level: Product formed: 30mm fibre Structure formed: zigzag or solenoid structure How is the structure formed? ● Nucleosome associate with each other, further shortening the molecule 3rd level: Product formed: 300 nm fibre Proteins involved: protein scaffold How is the product formed? ● The 30 nm chromatin fibre formeds looped domains attached to a protein scaffold Final level: Products formed: 700 nm metaphase chromatid How is this product formed? ● During mitosis/meiosis, the 300 nm fibre coil and fold,further compacting the chromosomes REGULATION IN CHROMATIN LEVEL ➔ Tightening or loosening of chromatins nucleosomes ( refer to dna packing) in order to increase or decrease rate of transcription ➔ Undergoes covalent modification by modifying such as acetylation methylation and phosphorylation

Histone modification Enzyme involved: Histone acetyltransferase (HAT) and Histone deacetylase (HDAC) Loosening of chromatin structure (histone acytelation) Enzyme: HAT What does HAT do? ● Catalyses the addition of acetyl group to lysine side chains ● Neutralises positive charge on lysine side chains and disrupts electrostatic attraction between histone protein and negatively charged DNA backbone ( phosphate group) ● As the forces are disrupted, chromatin structure is loosened, allows GTF to AND RNA Polymerase to access prometer more readily, INCREASING RATE OF TRANSCRIPTION Tightening of chromatin structure (histone deactyelasation