Protein Synthesis Lecture 12 & 13

Protein Synthesis Sections8.1 Protein Synthesis Learning Objectives Explain the role of tRNAs in translation Describe the structure and function of ribosomes Contrast the initiation of translation in bacterial and eukaryotic cells Outline the events of initiation, elongation, a...

Protein Synthesis Sections8.1 Protein Synthesis Learning Objectives Explain the role of tRNAs in translation Describe the structure and function of ribosomes Contrast the initiation of translation in bacterial and eukaryotic cells Outline the events of initiation, elongation, and termination of translation Summarize the mechanisms that regulate translation of specific mRNAs and global translation rates Protein Synthesis Key Concepts tRNAs serve as adaptors that align amino acids on the mRNA template. Peptide bond formation is catalyzed by rRNA in the ribosome. Translation initiation is different in prokaryotes and eukaryotes due to the lack of a nucleus in prokaryotes. In both cases, the ribosomal small subunit and initiator tRNA first interact with the mRNA before the large ribosomal subunit joins the complex. Translation begins at a start codon and ends at a stop codon. These are specific sequences found in the mRNA. Translation of mRNAs can be regulated by repressor proteins and miRNAs Global translational activity can be regulated by modification of translation initiation factors. Translation is literally switching languages! *Can you predict the anticodon sequence on the tRNA corresponding to a given Image source: Khan Academy codon? Ribosomes are made of structural _________ and catalytic _____ Overview of translation: three key stages Translation initiation signals differ in eukaryotes and prokaryotes Prokaryotic: Eukaryotic: Prokaryotic mRNAs can be ___________________  NOTE CHECK Which of the following is NOT a functional implication of the difference between prokaryotic and eukaryotic RNA processing and ribosome recognition strategies? a) Prokaryotic translation can begin while genes are still in the process of being transcribed b) Prokaryotic cells could not translate eukaryotic transcripts, and vice versa c) Multiple genes/proteins in a related process can be controlled under a single eukaryotic promoter NOTE CHECK Join.nearpod.com pin = Eukaryotic Initiation: eIFs – eukaryotic __________________ Small Subunit + eIFs + GTP-bound initiator tRNA mRNA + eIFs recognize 5’ cap and 3’ poly-A tail SSU complex loads onto mRNA at 5’ cap and scans along mRNA for ____________, powered by ATP. Hydrolysis of GTP bound to eIFs releases initiation factors. Binding of _________________ to form initiation complex. Elongation uses E____, P________, and A______________ sites  NOTE CHECK Which of the large ribosomal subunit sites may contain an uncharged tRNA, even if it is only temporary? a) A site only b) P site only c) E site only NOTE CHECK d) A & P sites e) P & E sites f) A, P, & E sites Join.nearpod.com pin = Termination via ______________ factors and _______ codon Translation Summary  NOTE CHECK Which of the following correctly aligns the orientation of the DNA template, mRNA transcript, and translated protein? a) 5’ -----DNA----- 3’ 3’ -----RNA----- 5’ N ---protein----C b) 3’ -----DNA----- 5’ NOTE CHECK 5’ -----RNA----- 3’ N ---protein----C c) 5’ -----DNA----- 3’ 5’ -----RNA----- 3’ C ---protein---- N Join.nearpod.com pin = The Central Dogma, but in reality rRNA Polypeptide → Functional Protein m tRNA Translated polypeptides are not automatically functional proteins Section 8.3 Lipid Chaperones modification Enzymes Disulfide bonds Modified from Wang et al. 2013  How do we control all of this? Transcriptional, Post-Transcriptional, and Post-Translational Regulation Sections 7.1, 7.3 Gene Regulation in E. coli Learning Objectives (Section 7.1) Explain how lactose regulates transcription of the lac operon Distinguish between positive and negative control Explain why repressors inhibit but activators stimulate transcription Key Concepts The lac operon is a model of gene regulation in bacteria although there are many other operons that regulate gene expression through positive and negative mechanisms The lac operon is negatively regulated by a constitutively expressed repressor protein that binds to the operator sequence and blocks RNA polymerase binding. The lac operon is under positive regulation by the binding of cAMP-bound CAP upstream of the promoter and facilitates the recruitment of RNA polymerase to the promoter resulting in transcriptional activation. Metabolism of lactose, an optional nutrient for E. coli, is a classic example of transcriptional regulation Sugars present in What E. coli want to environment metabolize Glucose only Lactose only Glucose and Lactose So… under what conditions does E. coli need to transcribe enzymes to metabolize lactose? Negative control of the lac ____________ Structural Genes: z y a Regulatory Control: P o i Transcription ______ unless _________ present Positive control of the lac operon by ____ is regulated by _______ Glucose 𝛼-ketoglutarate Transcription ______ unless glucose ___________  Note Check Imagine a bacterial line in which there was a deletion of 10 critical bases in the operator sequence. If glucose and lactose levels are both high, would LacZ be transcribed? a) Yes, because the Lac repressor would be unable to bind to the operator region due to the deletion. b) No, because the Lac repressor would be bound to the operator region due to the high levels of lactose. c) No, because cAMP would not be bound to CAP, so even without repressor binding there would be no activation. NOTE CHECK Join.nearpod.com pin =  Note Check Suppose there was a mutation in the repressor protein that caused allolactose to bind irreversibly to the repressor (once bound, it cannot be removed from repressor). After a period where lactose levels were high, you switch the cells to an environment with low levels of both glucose and lactose. Would you expect LacZ to be expressed? a) Yes, because the repressor would remain unable to bind to the operator even without high levels of lactose in the environment. b) No, because lactose levels are low so the repressor would be bound to the operator. c) No, because without glucose present, cAMP would not bind CAP. NOTE CHECK Join.nearpod.com pin = Chromatin and Epigenetics (Section 7.3) Describe the effects of different histone modifications on transcription Summarize the action of chromatin remodeling factors Key Concepts Enzymes that catalyze histone acetylation are associated with transcriptional activators. Enzymes that remove acetyl groups are associated with transcriptional repressors. Specific histone modification patterns are characteristic of transcriptionally active and inactive chromatin. DNA is packaged into _________________ DNA is wrapped around proteins called _____________ to form ___________________, the basic structural unit of chromatin What types of amino acids would you expect to find on the surface of histone proteins?  (euchromatin) (heterochromatin) Two chromatin states Euchromatin Heterochromatin Histones can undergo ___________ to promote open euchromatin Transcriptional activators and repressors associate with histone modifying enzymes Activators associate with Repressors associate with _________________________ (HATs) _________________________ (HDACs) which _______ acetyl groups to which _______ acetyl groups to __________ transcription __________ transcription ______________ and ______________ also modify histones Note: DNA itself is also modified directly by addition of methyl groups to cytosine residues!  Note Check Gene X is expressed in the brain, but not in the muscle. Which would you expect to be true? a) Gene X is likely absent from the genome of muscle cells b) Histones associated with Gene X are likely acetylated in the muscle, but not in the brain. c) Histones associated with Gene X are likely H3K4me in the brain, but H3K9me in the muscle. NOTE CHECK Join.nearpod.com pin = Transcriptional, Post-Transcriptional Regulation, and Post-Translational Regulation