Gene ExpressionPath from a gene to a Gene expression occurs in two steps: phenotype Transcription“Central Dogma of Copies information from a Molecular Biology” DNA sequence (a gene) to a complementary RNA sequenceDNA RNA Protein Translation Converts RNA sequence to amino acid sequence of a polypeptide
RNARNA (ribonucleic acid) differs from DNA:• Usually one polynucleotide strand• The sugar is ribose• Contains uracil (U) instead of thymine (T)
RNABases in RNA can pair with a single strand of DNA, except that adenine pairs with uracil instead of thymine.Single-strand RNA can fold into complex shapes by internal base pairing.
Three Types of RNAMessenger RNA (mRNA) Carries copy of a DNA sequence to site of protein synthesis at the ribosomeTransfer RNA (tRNA) Carries amino acids for polypeptide assemblyRibosomal RNA (rRNA) Catalyzes peptide bonds and provides structure
Transcription (DNA RNA)Transcription components:A DNA template for base pairings (one of the two strands of DNA)Nucleotides (ATP,GTP,CTP,UTP) as substratesRNA polymerase enzyme
RNA Polymerase• Catalyze the Synthesis of RNA• Do NOT need a primer to start synthesizing RNA• Do NOT have proofreading ability
Phases of TranscriptionInitiationElongationTermination
InitiationRNA Polymerase binds to the target DNA at the promoterUnwinds a ~10bp bubble of the DNA to initiate transcriptionTranscription begins at the initiation site
PromoterSequence of DNA where RNA polymerase bindsCritical Regulation Point for the Flow of Genetic Information in the CellTells RNA polymerase “When, Where, How Much”
ElongationRNA polymerase unwinds DNA about ten base pairs at a time; reads template in 3′ to 5′ direction.The RNA transcript is antiparallel to the DNA template strand, and adds nucleotides to its 3′ end.RNA polymerases do not proofread and correct mistakes.
TerminationIs specified by a specific DNA base sequence (termination site).Mechanisms of termination are complex and varied.For some genes the transcript falls away from the DNA template and RNA polymerase (others require another protein to assist)
Following TranscriptionThe completed mRNA leaves the nucleus through the nuclear poreMust interact with a ribosome to initiate translationInformation encoded on the mRNA is the utilized to guide protein synthesis
Genetic CodeSpecifies which amino acids will be used to build a proteinCodon:A sequence of three bases—each codon specifies a particular amino acid.Start codon: AUGInitiation signal for translation.Stop codons: UAA, UAG, UGAStop translation and polypeptide is released.
Genetic Code20 amino acids64 different possible codonsCodons are redundantGenetic Code is Universal
tRNAsTransfer RNAs act as the adapter between the mRNA and the growing polypeptideMust be “charged” or bound to a specific amino acid before they can be used in translation
tRNAsClover leaf secondary structureTwo important sites:Anticodon Binds to mRNAAmino Acid Attachment Site Binds to the respective amino acid
RibosomeThe workbench that holds mRNA and charged tRNAs in the correct positions to allow assembly of polypeptide chain.Ribosomes are not specific, they can make any type of protein.Composed of a Large and Small Subunit
Large SubunitThree tRNA binding sites:A (amino acid) site binds with anticodon of charged tRNAP (polypeptide) site is where tRNA adds its amino acid to the growing chainE (exit) site is where tRNA sits before being released from the ribosome.
Small SubunitHas a fidelity function: When proper binding occurs, hydrogen bonds form between the base pairs.
Phases of Translation• Initiation• Elongation• Termination
InitiationInitiation Complex Forms1) mRNA2) Charged tRNA (bound to methionine)3) Small subunit bound to the mRNA at the start codon
InitiationThe large subunit then joins the complex;The charged tRNA is now in the P site of the large subunit.
ElongationThe second charged tRNA enters the A site.Large subunit catalyzes two reactions:• It breaks bond between tRNA in P site and its amino acid• Peptide bond forms between that amino acid and the amino acid on tRNA in the A site
Elongation• The large subunit has peptidyl transferase activity• Critical to growing the polypeptide chain
ElongationWhen the first tRNA has released its methionine, it moves to the E site and dissociates from the ribosome—can then become charged again.Elongation occurs as the steps are repeated, assisted by proteins called elongation factors.
TerminationTranslation ends when a stop codon enters the A site.Stop codon binds a protein release factor— allows hydrolysis of bond between polypeptide chain and tRNA on the P site.Polypeptide chain separates from the
Post-Translational ModificationsProteolysis: Cutting of a long polypeptide chain into final products, by proteasesGlycosylation: Addition of sugars to form glycoproteinsPhosphorylation: Addition of phosphate groups catalyzed by protein kinases— charged phosphate groups change the conformationLeads to a mature and functional protein