Exam 3 Review Sheet Ch 14 15 16 17 Ch 14 DNA synthesis after Harvey Chase expt How does Replication get started origin of replication replication bubble forms in a chromosome when DNA is actively being synthesized Grows as replication proceeds bc synthesis is bidirectional occurs at both directions at same time replication fork Y shaped region where parent DNA double helix is split into 2 single strands which are then copied How is the helix opened and stabilized 1 Helicase enzyme breaks H bonds between deoxyribonucleotides 2 strands of DNA separate 2 single strand DNA binding proteins SSBPs attach to separated strands and prevent reformation of the double helix 3 topoisomerase cuts DNA to relieve twisting stress induced by helicase DNA helix is now open and stabilized How is the leading strand synthesized Recall that DNApolymeraseIII only works in 5 3 direction Synthesis requires a 3 end and a single stranded template 1 DNApolyIII requires primer a few nucleotides bonded to template strand to provide free 3 OH group to combine with incoming dNTP to form phosphodiester bond This is done by primase a type of RNApoly that catalyzes the polymerization of ribonucleotides into RNA 2 Once primer is in place DNApolyIII adds dNTPS to 3 end producing new strand with sequence complementary to template 3 As DNApolyIII moves along DNA sliding clamp holds the enzyme in place How is the Lagging strand synthesized DNApolyIII must work away from the replication fork to synthesize in 5 3 direction It lags behind synthesis occurring in the fork 1 Primase synthesizes short stretch of RNA that acts as primer DNApolyIII adds bases to the 3 end of the primer Enzyme moves away from replication fork but helicase continues to open the fork to expose new ssDNA on lagging strand 2 primase adds primer to newly exposed ssDNA DNApolyIII synthesizes lagging strand until it reaches fragment produced earlier Okazaki fragments 1000 bp long 3 DNApolyI removes primer replaces with dNTPS in 5 3 direction 4 DNA ligase catalyzes phosphodiester bond between adjacent fragments What happens at the end of linear chromosomes The lagging strand is too short because DNA synthesis cannot occur after primer is removed Telomerase enzyme that catalyzes synthesis of DNA from a built in RNA template 1 Unreplicated segment of telomere at 3 lagging strand forms a ss overhang 2 Telomerase binds to overhanging section of ssDNA Adds deoxyribonucleotides in 5 3 direction that is complementary to its RNA template 3 Primase DNApoly and ligase then synthesize in 5 3 direction to prevent chromosome shortening How does DNA Polymerase correct for mistakes 1 2 3 4 Error detection Nucleotide excision Nucleotide replacement by DNApoly in 5 3 direction Nucleotide linkage by DNA ligase Ch 15 Genetic Code For the following statements indicate whether they describe a Phenotype b Genotype A cat has long black hair A cat with long hair is encoded by genes H H A mutation in H gives rise to change in eye color Ch 16 Transcription RNA Processing Translation Eukaryotic RNA Polymerases Name of Enzyme RNA pol I RNA pol II RNA pol III Type of Gene Transcribed Large RNA molecules found in ribosomes Protein coding genes produce mRNA tRNAs How Does Transcription Begin Initiation phase 1 Sigma binds RNApol to form holoeynzme Sigma recognizes and binds to promoter regions of DNA Sigma is a regulatory protein In euks basal transcription factors instead of sigma Bacteria promoters 10 TTGACA and 35 box upstream from transcription start site Euk more diverse and complex TATA box at 30 from start How does Elongation and Termination Occur RNApol moves along DNA template in 3 5 RNA synthesis occurs in 5 3 direction Rudder steers template and nontemplate strands through channel inside enzyme Zipper helps separate newly synthesized RNA from DNA template Termination when RNA pol reaches DNA sequence that functions as transcription termination signal bacteria RNA folds back on itself to form hairpin Comparing Transcription in Bacteria and Euks RNA pol Promoter Proteins involved in contacting promoter RNA processing Translation Bacteria 1 35 10 box Sigma different versions of sigma bind different promoters None Euks 3 Variable TATA at 30 Basal transcription factors 1 5 cap 2 Splicing of introns 3 3 polyA tail tRNA adaptor molecule between mRNA and amino acids ribosomes catalytic machinery mRNA message to be translated amino acids building blocks of proteins ATP GTP potential energy What does a tRNA look like short RNA sequence 75 86 bp single stranded form secondary structures H bond with sequences in same molecules such as stemp loop 3 CCA is binding site for amino acids triplet on far end loop serves as anticodon Anticodon base pairs with mRNA codon How to initiate Translation AUG start codon initiation common in bacteria and euks 1 Ribosome binding site on mRNA binds to complementary sequence in small subunit of ribosome 6 nucleotides upstream of AUG Mediated by Initiation factors 2 Initiator Aminoacyl tRNA binds AUG f Met in bacteria normal methionine in euks 3 Large subunit joins to complete the complex tRNA is in P site How is the polypeptide extended Elongation Peptide bond formation the essence of protein synthesis occurs when P and A site are occupied by tRNAs 1 Arrival of animoacyl tRNA 2 Peptide bond forms 3 Translocation when elongation factors move mRNA through the ribosome in 5 3 direction Requires GTP How is translation terminated When translocation opens the A site and exposes a STOP codon a release factor protein fills the A site Hydrolysis of A site frees the polypeptide in the P site ribosome subunits separate
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