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Chemistry & Biochemistry 142C/242C Perm #________________________ Second Midterm May, 12th, 2009 These point assignments are not definitive of the final point values. They have to be considered as only estimate of the final point values. Questions 1 2 3 4 5 6 7 8 9 10 11 12 Points/Possible /5 /7 /5 /6 /5 /12 /8 /10 /14 /5 /7 /21 _______________ TOTAL /100 1) Draw the mechanism of the first step that takes place during the charging of a tRNA with its cognate amino acid. Identify the essential reactants and the product created in this reaction. (5 Points) 2) Explain how the isoleucyl-tRNA synthetase is able to distinguish between isoleucine and valine. Given the structure of histidine, propose a reason why histidyl-tRNA synthetase lacks a proofreading function. (7 points) Isoleucine is similar in structure to several other amino acids, particularly valine. Distinguishing between valine and isoleucine in the aminoacylation process requires the second filter of a proofreading function. Valine is able to fit into a second catalytic pocket where hydrolysis and release of the amino acid occurs. Histidine has a structure unlike that of any other amino acid, and this structure provides opportunities for binding specificity adequate enough to ensure accurate aminoacylation of the cognate tRNA. 3) Methionine is one of two amino acids with only one codon. How does the single codon for methionine specify both the initiating residue and interior Met residues of polypeptides synthesized by E. coli. (5 Points) There are two tRNAs for methionine: tRNAfMet, which is the initiating tRNA, and tRNAMet, which can insert a Met residue in interior positions in a polypeptide. Only fMet-tRNAfMet is recognized by the initiation factor IF-2 and is aligned with the initiating AUG positioned at the ribosomal P site in the initiation complex. AUG codons in the interior of the mRNA can bind and incorporate only Met- tRNAMet. 4) Briefly describe the sequence of events that take place during the initiation step of protein synthesis. (6 Points) Initiation factors 1 and 3 bind to the 30S subunit, which allows for binding to the mRNA at the Shine Dalgrano sequence, thus aligning the mRNA-rRNA interaction. IF1 blocks A-site and IF3 prevents preamture 50S association. With the help of IF2GTP, fMet-tRNAfMet then binds to the initiation codon AUG at the P site. Lastly, the 50S subunit of the ribosome is able to recognize the 30S complex. Upon binding the initiation factors are released. 5) The specific sequences that E. coli RNA polymerase usually binds to in E. coli DNA before initiating transcription generally contain more A=T base pairs than GC base pairs. Why this might be the case? (5pts) Because A=T base pairs are stabilized by only two hydrogen bonds (compared with three for GC pairs), double-stranded regions rich in A=T pairs are easier for RNA polymerase to bind and unwind in preparation for the transcription of one of the DNA strands. 6) What do you find on the 3' and 5' ends of eukaryotic mRNA after processing?(3 pts) Draw the chemical structure of the 5'end of an eukaryotic mRNA after it has been processed. (12 pts) At the 5' end there is a 7' methylguanosine cap consisting of a guanosine joined to the 5'-terminal nucleotide through a 5' to 5' triphosphate group. This guanine nucleotide is methylated on N-7. The next two nucleotides in the chain are also sometimes methylated on their 2'-OH groups. At the 3' end is the poly(A) tail consisting of a run of 80250 adenylate residues. 7) Compare transcription and reverse transcription in terms of the following characteristics. Fill in the table below. (8pts) Reverse Transcription (a) direction of polynucleotide synthesis (b) nature of template (c) nature of primer (d) incorporated nucleotides 5' to 3' RNA or DNA tRNA dNTPs Transcription 5' to 3' DNA none NTPs 8) Indicate whether each of the following statements is true (T) or false (F). (10 Points)
_F__Assembly of a complete ribosome onto an mRNA requires ATP hydrolysis. _T__Aminoacylation or "charging" of tRNA requires the formation of an aminoacylAMP intermediate. _F__Aminoacyl-tRNA binding to the A site of the ribosome requires the accessory factor EF-G and GTP hydrolysis. _T__Translocation of a growing polypeptide from the A to the P site on the ribosome requires EF-G and GTP hydrolysis. _F__Termination of translation requires release factors, but no NTP hydrolysis. _T__ Bacterial mRNA is broken down within a few minutes of its formation in E. coli. _F__ Bacterial mRNA consists only of the bases that code for amino acids. _F__ Polysomes do not contain mRNA. _F__ Bacterial mRNA normally occurs as a double-stranded structure, with one strand containing codons, the other containing anticodons. _T__ Bacterial mRNA can be translated while it is still being synthesized. 9) What type of chemical reactions are catalyzed by group II introns? Draw the chemical reaction
of the first step of splicing catalyzed by group II intron. (Draw the detailed chemical structure of the nucleophile and splice site, indicate the mechanism of the reaction by arrows and draw the final product). (14 pts) Transesterification reaction (see reaction for group I intron for detailed view) 2' OH of adenine attacks Phosphate. 10) What is the purpose of posttranslational modification of amino acids in a newly synthesized polypeptide chain? Name two types of modifications and how they affect the protein. (5 Points) Posttranslational modifications are made to a newly synthesized polypeptide so the protein can attain its biologically active conformation . Examples of posttranslational modifications and their affects are on page 1097 in Lehninger. 11) Describe how both prokaryotes and eukaryotes determine the correct reading frame during protein synthesis (7 Points). Prokaryotes: Recognition of the Shine-Dalgarno sequence aligns the ribosome in the correct reading frame. Eukaryotes: The initiation complex (eIF4F) ties together the 5' and 3' end of the mRNA. The ribosome then scans the mRNA and the first (5')AUG sequence that is encountered signals the beginning of the reading frame. 12) MULITPLE CHOICE (1 POINT EACH) After binding by E. coli RNA polymerase, the correct order of events for transcription initiation is: A) closed complex formation, open complex formation, promoter clearance, start of RNA synthesis. B) closed complex formation, open complex formation, start of RNA synthesis, promoter clearance. C) open complex formation, closed complex formation, start of RNA synthesis, promoter clearance. D) start of RNA synthesis, closed complex formation, open complex formation, promoter clearance. E) start of RNA synthesis, open complex formation, closed complex formation, promoter clearance. Processing of a primary mRNA transcript in a eukaryotic cell does not normally involve: A) attachment of a long poly(A) sequence at the 3' end. B) conversion of normal bases to modified bases, such as inosine and pseudouridine. C) excision of intervening sequences (introns). D) joining of exons. E) methylation of one or more guanine nucleotides at the 5' end. Which one of the following is true about the genetic code? A) B) C) D) All codons recognized by a given tRNA encode different amino acids. It is absolutely identical in all living things. Several different codons may encode the same amino acid. The base in the middle position of the tRNA anticodon sometimes permits "wobble" base pairing with 2 or 3 different codons. E) The first position of the tRNA anticodon is always adenosine. The excision (splicing) of many group I introns requires, in addition to the primary transcript RNA: A) B) C) D) E) a cytosine nucleoside or nucleotide and a protein enzyme. a guanine nucleoside or nucleotide (only). a protein enzyme only. a small nuclear RNA and a protein enzyme. ATP, NAD, and a protein enzyme. Which of the following is not true of tRNA molecules? A) B) C) D) E) The 3'-terminal sequence is --CCA. Their anticodons are complementary to the triplet codon in the mRNA. They contain more than four different bases. They contain several short regions of double helix. With the right enzyme, any given tRNA molecule will accept any of the 20 amino acids. In E. coli, aminoacyl-tRNA synthetases: A) activate amino acids in 12 steps. B) are amino acidspecific; there is at least one enzyme specific for each amino acid. C) fall into two classes, each of which attaches amino acids to different ends of the tRNA. D) have no proofreading activities. E) require a tRNA, an amino acid, and GTP as substrates. The enzyme that attaches an amino acid to a tRNA (aminoacyl-tRNA synthetase): A) B) C) D) E) always recognizes only one specific tRNA. attaches a specific amino acid to any available tRNA species. attaches the amino acid at the 5' end of the tRNA. catalyzes formation of an ester bond. splits ATP to ADP +Pi. A branched ("lariat") structure is formed during: A) B) C) D) E) attachment of a 5' cap to mRNA. attachment of poly(A) tails to mRNA. processing of preribosomal RNA. splicing of all classes of introns. splicing of group II introns. Differential RNA processing may result in: A) B) C) D) E) a shift in the ratio of mRNA produced from two adjacent genes. attachment of the poly(A) tail to the 5' end of an mRNA. inversion of certain exons in the final mRNA. the production of the same protein from two different genes. the production of two distinct proteins from a single gene. Reverse transcriptase: A) can utilize only RNA templates. B) has a 3' 5' proofreading exonuclease but not a 5' 3' exonuclease. C) is activated by AZT. D) is encoded by retroviruses. E) synthesizes DNA with the same fidelity as a typical DNA polymerase. Which one of the following statements about mRNA stability is true? A) Degradation always proceeds in the 5' to 3' direction. B) Degradation of mRNA by polynucleotide phosphorylase yields 5'-nucleoside monophosphates. C) In general, bacterial mRNAs have longer half-lives than do eukaryotic mRNAs. D) Rates of mRNA degradation are always at least 10-fold slower than rates of mRNA synthesis. E) Secondary structure in mRNA (hairpins, for example) slows the rate of degradation. Which of the following statements about the synthesis of rRNA and tRNA in E. coli is true? A) Both rRNA and some tRNAs are part of the same primary transcript. B) Each rRNA sequence (16S, 23S, 5S) is transcribed into a separate primary transcript. C) Primary tRNA transcripts undergo methylation, but rRNA sequences are not methylated. D) The tRNA sequences all lie at the 3'end of the rRNA transcripts E) There is a single copy of the rRNA genes. Compared with DNA polymerase, reverse transcriptase: A) B) C) D) does not require a primer to initiate synthesis. introduces no errors into genetic material because it synthesizes RNA, not DNA. makes fewer errors in synthesizing a complementary polynucleotide. makes more errors because it lacks the 3' 5' proofreading exonuclease activity. E) synthesizes complementary strands in the opposite direction from 3' to 5' Which of the following is true about the sorting pathway for proteins destined for incorporation into lysosomes or the plasma membrane of eukaryotic cells? A) Binding of SRP to the signal peptide and the ribosome temporarily accelerates protein synthesis. B) The newly synthesized polypeptides include a signal peptide at their carboxyl termini. C) The signal peptide is cleaved off inside the mitochondria by signal peptidase. D) The signal recognition particle (SRP) binds to the signal peptide soon after it appears outside the ribosome. E) The signal sequence is added to the polypeptide in a posttranslational modification reaction. ...
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This note was uploaded on 07/13/2009 for the course CHEM 142c taught by Professor Reich,n during the Spring '08 term at UCSB.
- Spring '08