B1115 - SP24 - Stud - Ch 17 - Transcription, RNA Processing, and Translation.pdf

Transcription, RNA Processing, and Translation Why Are Proteins Important? List at least three types of proteins that we have talked about so far in this course. Explain the function of each of these types of proteins. Which of the following correctly indicates the complementary base pairing of aden...

Transcription, RNA Processing, and Translation Why Are Proteins Important? List at least three types of proteins that we have talked about so far in this course. Explain the function of each of these types of proteins. Which of the following correctly indicates the complementary base pairing of adenine in DNA and RNA? A. B. C. D. E. adenine with cytosine in DNA and with guanine in RNA adenine with guanine in DNA and with cytosine in RNA adenine with thymine in both DNA and RNA adenine with thymine in DNA and with uracil in RNA adenine with uracil in DNA and with thymine in DNA Which of the following correctly indicates the complementary base pairing of adenine in DNA and RNA? A. B. C. D. E. adenine with cytosine in DNA and with guanine in RNA adenine with guanine in DNA and with cytosine in RNA adenine with thymine in both DNA and RNA adenine with thymine in DNA and with uracil in RNA adenine with uracil in DNA and with thymine in DNA The information carried by a DNA molecule is in: A. its amino acid sequence. B. the sugars and phosphates forming its backbone. C. the order of the bases in the molecule. D. the total number of nucleotides it contains. E. the RNA units that make up the molecule. The information carried by a DNA molecule is in: A. its amino acid sequence. B. the sugars and phosphates forming its backbone. C. the order of the bases in the molecule. D. the total number of nucleotides it contains. E. the RNA units that make up the molecule. Lecture Outline 1. Transcription 2. RNA Processing 3. Translation a) tRNAs b) Ribosomes c) Process of Translation 4. Post-Translational Modifications Lecture Outline 1. Transcription 2. RNA Processing 3. Translation a) tRNAs b) Ribosomes c) Process of Translation 4. Post-Translational Modifications Overview of Transcription RNA polymerases synthesize an mRNA version of the instructions stored in DNA. Only one strand of DNA, called the template strand, is copied. Other strand is the non-template, or coding strand. Transcription As a region of DNA unwinds, one strand is used as a template for the RNA transcript to be made. Nucleoside triphosphates (NTPs) are used to produce an RNA transcript that is complementary to the template strand. The new strand grows in the 5’à 3’ direction. Which four NTPs are used for RNA synthesis? ATP, GTP, CTP, UTP In which direction is the DNA template strand read? 3’ to 5’ Transcription: Initiation RNA polymerase and associated proteins bind to the DNA duplex at promoter sequences. o These regions contain sequences that indicate where a gene starts and which DNA strand is the template strand. o Promoters are located upstream of the transcription start site. Transcription RNA polymerase opens the DNA double helix, creating a transcription bubble. The template strand is threaded through the RNA polymerase active site. Incoming NTPs enter a channel in the enzyme and diffuse to the active site. NTPs pair with complementary DNA bases, and polymerization begins. Transcription: Elongation Transcription: Termination Transcription ends with termination. RNA polymerase transcribes a transcription-termination signal. In prokaryotes, this codes for RNA that forms a hairpin structure. Causes the RNA polymerase to separate from the RNA transcript. In eukaryotes, a poly(A) signal is transcribed rather than a hairpin, and the RNA downstream is cut. Which of the following correctly lists the components necessary for prokaryotic and eukaryotic transcription? A. DNA polymerase, general transcription factors, DNA, and DNA nucleotides B. ribosomes, general transcription factors, DNA, and DNA nucleotides C. ribosomes, general transcription factors, DNA, and RNA nucleotides D. RNA polymerase, general transcription factors, DNA, and DNA nucleotides E. RNA polymerase, general transcription factors, DNA, and RNA nucleotides Which of the following correctly lists the components necessary for prokaryotic and eukaryotic transcription? A. DNA polymerase, general transcription factors, DNA, and DNA nucleotides B. ribosomes, general transcription factors, DNA, and DNA nucleotides C. ribosomes, general transcription factors, DNA, and RNA nucleotides D. RNA polymerase, general transcription factors, DNA, and DNA nucleotides E. RNA polymerase, general transcription factors, DNA, and RNA nucleotides RNA nucleotides are always added to the 3’end of a growing RNA polynucleotide. A. True B. False RNA nucleotides are always added to the 3’end of a growing RNA polynucleotide. A. True B. False Lecture Outline 1. Transcription 2. RNA Processing 3. Translation a) tRNAs b) Ribosomes c) Process of Translation 4. Post-Translational Modifications Primary Transcript vs. mRNA The RNA transcript that comes off the template DNA strand is known as the primary transcript. It contains the information of the gene that was transcribed. For protein-coding genes, the primary transcript includes the information needed to direct the ribosome to produce the protein. – The RNA molecule that combines with the ribosome to direct protein synthesis is called mRNA. Primary Transcript in Prokaryotes In prokaryotes, this relationship is simple because the primary transcript is the mRNA. Both processes occur in the cytoplasm and there is no nuclear envelope to spatially separate transcription from translation. Primary Transcript in Eukaryotes In eukaryotes, there is a barrier between transcription and translation (the nuclear membrane). The primary transcript undergoes a complex process of chemical modifications known as RNA processing. Three types of chemical modification occur before the mRNA is translated by the ribosome: RNA splicing Addition of a 5’ cap Polyadenylation RNA Splicing One modification of the primary transcript is the excision of certain sequences known as introns, leaving intact the exons. This process is known as RNA splicing. About 90% of all human genes contain at least one intron. RNA Splicing Splicing allows different mRNAs and proteins to be produced from a single gene. One primary transcript can code for multiple proteins; which protein is formed depends on how the transcript is spliced. Such variations are called alternative RNA splicing. Adding Caps and Tails to Transcripts Primary RNA transcripts are also processed by the addition of: A 5¢ cap: a modified guanine nucleotide that enables ribosomes to bind and protects from degradation. A 3′ poly(A) tail: 100–250 adenine nucleotides; is needed for translation and protects from degradation. After splicing and addition of the cap and tail, the product is a mature mRNA. Mature mRNAs contain untranslated regions (UTRs) at both ends. An intron is ______________. A. a protein that is clipped out post-translationally B. a series of amino acids at the end of a new polypeptide that directs transcription to the ER C. a transfer RNA that binds to the codon D. part of an intact, mature mRNA that leaves the nucleus E. RNA that is removed during the processing of an RNA molecule and remains inside the nucleus An intron is ______________. A. a protein that is clipped out post-translationally B. a series of amino acids at the end of a new polypeptide that directs transcription to the ER C. a transfer RNA that binds to the codon D. part of an intact, mature mRNA that leaves the nucleus E. RNA that is removed during the processing of an RNA molecule and remains inside the nucleus Which of the following is not true of RNA processing? A. B. C. D. Exons are cut out before mRNA leaves the nucleus. Nucleotides may be added at both ends of the RNA. Ribozymes may function in RNA splicing. A primary transcript is often much longer than the final RNA molecule that leaves the nucleus. E. Introns are cut out before mRNA is translated. Which of the following is not true of RNA processing? A. B. C. D. Exons are cut out before mRNA leaves the nucleus. Nucleotides may be added at both ends of the RNA. Ribozymes may function in RNA splicing. A primary transcript is often much longer than the final RNA molecule that leaves the nucleus. E. Introns are cut out before mRNA is translated. Sometimes the sequence of DNA gets mutated and an adenine is paired to a cytosine. Why is this interaction unstable? A. Because an ionic bond cannot be formed between the two. B. Because adenine forms only two hydrogen bonds and not three. C. Because the phosphate groups of the two bases repel each other. D. Because the charges on the bases repel one another. Sometimes the sequence of DNA gets mutated and an adenine is paired to a cytosine. Why is this interaction unstable? A. Because an ionic bond cannot be formed between the two. B. Because adenine forms only two hydrogen bonds and not three. C. Because the phosphate groups of the two bases repel each other. D. Because the charges on the bases repel one another. How is it possible that an mRNA could have the correct sequence yet still not be translated appropriately? A. B. C. D. E. The 5’ cap was not added. The DNA sequence was incorrect. The polyA tail was not formed. a and c none of the above How is it possible that an mRNA could have the correct sequence yet still not be translated appropriately? A. B. C. D. E. The 5’ cap was not added. The DNA sequence was incorrect. The polyA tail was not formed. a and c none of the above Protein Synthesis During transcription, one of the two DNA strands called the template strand provides a template for ordering the sequence of nucleotides in an RNA transcript. During translation, the mRNA base triplets, called codons, are read in the 5¢ to 3¢ direction. Each codon specifies the amino acid to be placed at the corresponding position along a polypeptide. How many amino acids are there? Which amino acid does the codon AAC code for? A. Asparagine B. Glutamine C. Glycine D. Lysine E. Tryptophan Which amino acid does the codon AAC code for? A. Asparagine B. Glutamine C. Glycine D. Lysine E. Tryptophan Review List the three major types of RNA. mRNA, tRNA, rRNA Where do transcription and translation occur in prokaryotes? In the cytoplasm In eukaryotic cells? Transcription occurs in the nucleus. Translation occurs in the cytoplasm. Overview of Translation In translation, the sequence of bases in an mRNA is decoded to synthesize the amino acid sequence in a protein. Ribosomes catalyze translation of the mRNA sequence into protein. oIn many cases, polyribosomes will form where multiple ribosomes are attached to a single mRNA at a single time. § Many copies of a protein are produced from one mRNA. tRNAs bind to amino acids and then transfer them to the growing polypeptide. Location of Transcription and Translation In bacteria, ribosomes often begin translating an mRNA before transcription is complete. In eukaryotes, transcription and translation are separated. mRNAs are synthesized and processed in the nucleus. Mature mRNAs are transported to the cytoplasm for translation by ribosomes. Why do you think this is different in prokaryotes vs. eukaryotes? Lecture Outline 1. Transcription 2. RNA Processing 3. Translation a) tRNAs b) Ribosomes c) Process of Translation 4. Post-Translational Modifications What do tRNAs Look Like? tRNAs are relatively short: 75–85 nucleotides long. Flattened into one plane to reveal its base pairing, a tRNA molecule looks like a cloverleaf. Because of hydrogen bonds, tRNA actually twists and folds into a threedimensional molecule. tRNA is roughly L-shaped. What do tRNAs Look Like? A CCA sequence at the 3ʹ end is the binding site for amino acids. An aminoacyl tRNA is a tRNA linked to its amino acid. The loop at the opposite end forms the anticodon. Has a sequence of three nucleotides. Can base-pair with the mRNA codon. How Are Amino Acids Attached to tRNAs? Enzymes called aminoacyl-tRNA synthetases “charge” the tRNA by catalyzing the addition of amino acids to tRNAs. ATP is required to attach tRNA to an amino acid. For each of the 20 amino acids: There is a different aminoacyl tRNA synthetase. There are one or more tRNAs. How Many tRNAs Are There? There are 61 different codons but only about 40 tRNAs in most cells. Crick proposed the wobble hypothesis: The anticodon of tRNAs can still bind successfully to a codon whose third position requires a nonstandard base pairing. One tRNA is able to base-pair with more than one type of codon. How can this be possible without making mistakes during translation? Lecture Outline 1. Transcription 2. RNA Processing 3. Translation a) tRNAs b) Ribosomes c) Process of Translation 4. Post-Translational Modifications Ribosome Structure Ribosomes contain many proteins and ribosomal RNA (rRNA). Ribosomes can be separated into two subunits 1. The small subunit holds the mRNA in place. 2. The large subunit is where peptide bonds form. During translation three tRNAs can line up within the ribosome. Ribosome Structure The tRNAs can only fit when its anticodon binds to the corresponding codon in the mRNA. The tRNAs fit into three sites in the ribosome: 1. The A site is the acceptor site for an aminoacyl tRNA. 2. The P site is the peptidyl site where a peptide bond forms. 3. The E site is where tRNAs without amino acids exit the ribosome. Lecture Outline 1. Transcription 2. RNA Processing 3. Translation a) tRNAs b) Ribosomes c) Process of Translation 4. Post-Translational Modifications Translation The ribosome is a molecular machine that synthesizes proteins in a three-step sequence: 1. An aminoacyl tRNA carrying the correct anticodon for the mRNA codon enters the A site. 2. A peptide bond forms between the amino acid on the Asite tRNA and the polypeptide on the P-site tRNA. 3. The ribosome moves down the mRNA by one codon and all three tRNAs move down one position. The tRNA in the E site exits. The A site is available for another tRNA to bind. Translation The protein grows by one amino acid with each repeat of the three steps. Amino acids are always added to the carboxyl end (C-terminus) of the polypeptide. What is the C-terminus of a Translation has three phases: 1. Initiation 2. Elongation 3. Termination polypeptide? Translation: Initiation The initiation phase of translation begins near the AUG start codon. The small ribosomal subunit binds to the mRNA at the ribosome binding site. The ribosome binding site is about 6 bases upstream from the start codon. This process is mediated by initiation factors. The first tRNA is called the initiator tRNA. It carries a modified methionine. Translation: Initiation Translation initiation is a three-step process in bacteria: 1. The mRNA binds to a small ribosomal subunit. 2. The initiator tRNA bearing f-Met binds to the start codon. 3. The large ribosomal subunit binds so that the initiator tRNA is in the P site. Translation is now ready to begin. Translation: Elongation At the start of elongation: The initiator tRNA is in the P site. The E and A sites are empty. An aminoacyl tRNA binds to the codon in the A site. The amino acid on the P-site tRNA is connected to the amino acid on the A-site tRNA forming a peptide bond. Peptide Bond Formation Translation: Elongation Translocation occurs when the ribosome slides one codon toward the 3ʹ end of the mRNA. Elongation factors help move the ribosome. Translocation accomplishes three things: 1. The uncharged tRNA from the P site moves into the E site and is ejected from the ribosome. 2. The tRNA attached to the growing protein moves into the P site. 3. Opens the A site to expose a new codon, which is available to accept a new aminoacyl tRNA. Polypeptide Translation: Termination Termination occurs when the A site encounters a stop codon. A protein called a release factor enters the A site. Resembles tRNAs in size and shape. But does not carry an amino acid. Hydrolyzes the bond linking the P-site tRNA to the polypeptide chain. The newly synthesized polypeptide, tRNAs, and ribosomal subunits separate from the mRNA. Lecture Outline 1. Transcription 2. RNA Processing 3. Translation a) tRNAs b) Ribosomes c) Process of Translation 4. Post-Translational Modifications Post-Translational Modifications Most proteins go through an extensive series of processing steps called post-translational modification before they are completely functional. Folding determines a protein’s shape and function. Molecular chaperones speed protein folding. Which of the following parts does not take part in polypeptide synthesis? A. B. C. D. E. An exon mRNA An intron tRNA A ribosome Which of the following parts does not take part in polypeptide synthesis? A. B. C. D. E. An exon mRNA An intron tRNA A ribosome How does RNA polymerase ‘know’ where to start transcribing a gene into mRNA? A. It starts at one end of the chromosome. B. Transfer RNA acts to translate the message to RNA polymerase. C. It starts at a certain nucleotide sequence called a promoter. D. The ribosome directs it to the correct portion of the DNA molecule. E. It looks for the AUG start codon. How does RNA polymerase ‘know’ where to start transcribing a gene into mRNA? A. It starts at one end of the chromosome. B. Transfer RNA acts to translate the message to RNA polymerase. C. It starts at a certain nucleotide sequence called a promoter. D. The ribosome directs it to the correct portion of the DNA molecule. E. It looks for the AUG start codon.