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ch8 - STEP~BYaSTEP GUIDE Major Concepts Nucleotides have a...

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Unformatted text preview: STEP~BYaSTEP GUIDE Major Concepts Nucleotides have a variety of roles in cells. They serve as carriers of chemical energy, as compo- nents of enzyme cofactors, and as molecular messenv gers. Most important, they are the monomeric subunits of the polymers RNA and RNA. These nucieic acids carry the genetic information of every organism. Nucleic acids are polymers of nucleotide residues. Nucleotides contain three characteristic components: a nitrogenous base (purine or pyrimidine), a pentose (ribose or deoxyribose), and a phosphate group. The nucleotides in RNA contain ribose; in DNA, they core tain 2'—deoxyribose. ri‘he pyrimidine bases in RNA are uracil and cytosine, While in DNA they are thymine and cytosine. The purines are adenine and guardne in RNA and DNA. A nucleoside is a nitrogenous base covalently bound in an N-B-giycosidic linkage to the 1' carbon of a pentose sugar. A nucleotide is a nu- cleoside with at least one phosphate group attached to the 5’ carbon of the sugar. Successive nucleotide residues within a polymer are linked through their phosphate groups in phosphodiester linkages be» tween the 5’ hydroxyi of one pentose and the 3' hy- droxyl of the next. dust as individual amino acid residues influence the overall structure of the prom teins they make up, the chemical properties of the in- dividual nucleotide residues influence the structure and behavior of the nucleic acids they form. Nucleic acids have levels of structure, similar to proteins. Polymers of DNA or RNA exhibit polarity in that the two ends of the molecule are different. The 3’ end has no nucleotide attached at the 3' carbon of the pea tose, whereas the 5’ end has no nucleotide attached through the 5’ phosphate. Two polymers (strands) of DNA are associated in an antiparallel arrangement; the orientation of one strand (with respect to its 3’ 100 Nucleotides and Nucleic Acids and 5' ends) is opposite to that of the other strand. In the Watson-Crick DNA structure, also called B—form DNA, the strands are stabilized in a right—handed doua tile-helix conformation that is held together by hydro- gen bonds and base-stacking interactions. The ability of nucleotides in one strand to form specific base pairs with nucleotides in the other (adenine pairs with thymine; cytosine pairs with guanine) is the re» sult of hydrogen bonds. Consequently, each strand in a double helix complements the other with respect to nucleotide sequence. The overall content of the dif~ ferent bases can affect the form of the DNA. A—DNA is relatively shorter and thicker than B—DNA; Z—DNA is left—handed. The occurrence of structural variants may have biological significance. RNA is transcribed as a complementary, single— stranded molecule using a DNA molecule as a tem- plate. Three species of RNA in cells are messenger RNA (mRNA), ribosomai RNA (rRNA), and transfer RNA {tRNA). These different types of RNA have varv» ious secondary structures, which are related to their base sequence and, in part, determine their functions. An understanding of the chemistry of nucleic acids facilitates understanding of their functions. Disruption of the noncovalent hydrogen bonds and hydrophobic interactions between bases in the DNA double helix causes it to denature or “melt” into sep— arate strands. Laboratory measurements of the tem— perature at which a specific DNA denatures can yield information on its base composition (GEO pairs form more hydrogen bonds than do A=T pairs and there— fore lead to higher melting temperatures). Denat— uration of double-helical DNA is the first step in hybridization experiments that form the basis of many techniques used in molecular genetics. The _ - a 2% '"r a Step-By~Step Guide 101 sequence of a strand of DNA can be determined using What t0 Review automated procedures, and DNA polymers containing specific sequences can be chemically synthesized. Answering the following QHEStiOHS and reviewing the 1‘8?- evant concepts, which you have already studied, should DNA is susceptible to chemical changes that cause make this chapter more understandable. mutations. Deamination reactions, hydrolysis of N—B-glycosidic bonds, irradiation-induced formation of pyrimidine dimers, alkylating reactions, and oxidative damage all can cause mutations in DNA. Not all alterations to the N d 1 , . _ th 1 t' f giycosidic bonds (Figs. 7-~ll and 7—29). D. A are eeterious, enzymatic me ya 10“ O cer— - Nucleotides contain a carbohydrate residue in the tam bases is common and affects the structure and . . furanose form. Renew the structure and nomencla function of DNA. ture of furanoses. Draw a furanose sugar, including the numbering of its carbon skeleton (Fig. 7—7). ' In a nucleotide, the pentose sugar is linked to the purine or pyrimidine base through an N—fl—glycosidic bond. Review glycosidic bonds and remind yourself of other biologically important molecules that have Nucleotides serve important cellular functions oe— youd their role as information molecules. - Like proteins, nucleic acids exhibit several levels of Nucleoside triphosphates, most importantly ATP, structural organization Compare the levels of pro- transfer energy in the cell. Many cofactors, including tem structure (Chapters .3 and 4} _t0 the levels Of coenzyme A, contain nucleotides, as do second mes— “1101819 301d structure discussed m this chapter. sengers such as c AMP. What bonds and/or interactions are important to each class of biomolecule? - The nucleotide ATP affects the reaction equilib— rium and rate of many chemical reactions. Be sure you understand the distinction (pp. 186488). Topics for Discussion , Answering each of the following questions, especially in the context of a study group dis- cussion, should help you understand the important points of this chapter. 8.1 Some Basics 1. What are the functions of DNA and of the various types of RNA? 2. What is the functional defirution of a gene? Nucleotides and Nucleic Acids Have Characteristic Bases and Pcntcsss 3. What are the general constituents of nucleosides and nucleotides, and What types of bonds are involved in each of these structures? 4. Which bases are commonly found in DNA? Which bases are found in RNA? 5. What minor bases are occasionally found in DNA? In RNA? 6. What are the possible phosphorylation sites in nucleotides? 102 Chapter 8 Nucleotides and Nucleic Acids Phosphodiester Bands Link Successive Nucleotides in Nucleic Acids 7. 8. 9. What components form the backbone of DNA and RNA molecules? ls DNA soluble in water? Why or why not? Why is RNA sensitive to aikaline hydrolysis, whereas DNA is not? The Properties of Nucleotide Bases Affect the Three-Dimensional Structure of Nucleic Acids 10. 11. 8.2 Nucleic Acid Structure 12. How does pH affect the structure of the purine and pyrimidine bases? Are the tree bases soluble in water? What are the hierarchical levels of nucleic acid structure? What are the analogous lev- els of structure in proteins? DNA Is a Double Helix That Stores Genetic Information 7 13. 14. 15. 16. 17. Which of “Chargaff’s rules" do you think was the most important clue leading to the mode} postulated by Watson and Crick? Refer to Figure 8—14 and identify the three components that make up a nucleotide, the ribose sugar, the phosphate group, and the nitrogenous base. Which of these are re» sponsible for joining the nucleotides together in a chain and which are involved in the association of two nucleotide strands in a doubie helix? What type of noncovalent interaction is critical for the specificity of base pairing be- tween nucieotides of complementary strands of DNA? What types of noncovalent interactions contribute to the overall stability of the double heiix? Draw, as best you can in two dimensions, the structure of double—holiest DNA without iooking at your text! Be sure you know where each of the nucleotide components is placed in relationship to the others, and the bonds and interactions involved. How does this structure suggest a mechanism for the transmission of genetic information? DNA Can Occur in Different Three-Dimensional Farms 18. What are the similarities and differences among B, A-, and Z—DNA‘? Step-By-Step Guide 103 19. What is the possible biological significance of the alternate forms of DNA? Certain DNA Sequences Adapt Unusual Structures 20. What is the difference between a palindrome and a mirror repeat? 21. Why can’t mirror repeats form hairpin structures? 22. What chemical conditions promote the formation of triplex DNA strands? What nu- cieotide sequences promote triplex formation? 23. Why are these unusual DNA structures of interest? Messenger RNAs Code for Polypeptide Chains 242. What is the minimum length of mRNA required to code for a polypeptide chain of 80 amino acid residues? 25. What is one function of the noncoding regions of mRNA? Many RNAs Have More Complex Three-Dimensional Structures 26. How does base pairing differ in DNA—DNA interactions and RNA—RNA interactions? 27. What bonds and interactions are important in some RNA secondary structures? 28. What is it about RNA molecules that allows them to form more complex structures than do molecules of DNA? 8:;3 Nucleic Acid Chemistry Doublewefical DNA and RNA Can Be Denatured 29. Which bonds and interactions are affected during denaturation of DNA? How does this compare to protein denaturation? 30. What physical changes allow scientists to determine the melting points of DNA strands with different nucleotide compositions? How are these changes experimentally reea~ sured? 31. Remind yourself why guanine can form three hydrogen bonds with cytosine, Whereas aderunefithymine pairs have only two. 104 Chapter 8 Nucleotides and Nucleic Acids Nucleic Acids from Different Species Can Form Hybrids 32. How does the degree of evolutionary closeness between two species affect the extent of hybridization between their nucleic acids? Nucleotides and Nucleic Acids Undergo Nanenzymatic Transformations 33. Explain in evolutionary terms why it is important that DNA contains thymine rather than uracil. 34. Physicians caution against excessive sun tanning. Explain the molecular basis for their concern. Some Bases of DNA Are Methylated 35. List some reasons for methylation of certain DNA bases and sequences. The Sequences of Long DNA Strands Can Be Determined 36. What are the general principles involved in the two methods of sequencing DNA? 37. What is the roie of the dideoxynucleoside triphosphate in the Sanger method? 38. What component is labeled in the sequencing reactions using the Sanger method? 39. Be able to “read” a sequencing gel. The Chemical Synthesis of DNA Has Been Automated 40. The ability to synthesize DNA has proved to be an extremely useful tool for a wide va— riety of biochemical and biological studies. Think of a biochemical question that can be addressed using synthetic polynucieotides. 8.4 Other Functions of Nucleotides Nucleotides Carry Chemical Energy in Cells 41. Theoretically, how do ATP and PR compare in their ability to affect reaction equilibria? 42. How do ATP and PR compare in their ability to affect reaction rates? Adenine Nucleotides Are Components of Many Enzyme Cofactars 43. Explain the concept of evolutionary economy. How does it relate to the adenosine com- ponent of many cofactors? Step—By—Step Guide 105 Some Nucleotides Are Regulatory Molecules 44. Why is CAMP called a second messenger? Discussion Questions for Study Groups 0 Why is DNA so welinsoited to the role of molecular repository of genetic information and why is RNA not gener« ally used for this “information storage” function? 0 What are the cellular roles of the various types of RNA and how are prokazyotic and eukaryotic mRNA molecules different? 0 With improvements in automated DNA sequencing it Will someday become possible to obtain anyone’s complete DNA sequence. What are some possible benefits and problems associated with the ability to obtain your personal DNA sequence? 106 Chapter 8 Nucleotides and Nucleic Acids SELF-TEST Be You Know the Terms? IIIIIIHIIIIII HIIIIHIIIIII IIII I EIIII HIIIIIIIIIIIIIII ”E I “II IIIIIIIII W I x-n ya U! N h.) a E 8 I IIIIIIII M en E H M 01 WIIIII WIIIIIIII EIIIIIIIH ACROSS 15. A common protein domain found in proteins that bind ATP. (3 words) 1' These two strands are -—~——W to each ”the“ 16. A major structural component of the protein synthetic AATGCGGTCCTM‘ machinery of cells. TTACQCCAGGATA 18. Covalent bonds that fink the individual nucieotide 5' 3, ““9 5, residues in DNA and RNA. 5 (_ 3 _ , , ‘ ‘ . . ‘ 19. Structure containing polypurine tracts and mirror 7. A ribonucleic aczd involved in protein synthesm; it binds repeats forms a triple helix. ammoemds. _ _ 23. Left»handed double-helical structure. 3- contains a phOSPha‘e gm”? 11" a“ 93“” Wage to a 24. Right-handed, Watson-Crick doubie helix. 1t}. 12. 13. 14. ribose sugar A common intracelluiar signaling moiecule. (2 words) and a nitrogenous base. 25. Short polymers of nucleotides (50 or less), often used as complementary DNA “probes” for hybridization Most eukaryotic mRNA codes for a single poiypeptide techniques. and is ‘ 26. AACCT’I‘T’E‘GCAA What two complementary strands of DNA spontaneously TTGGE f 5 @GG'E‘T {2 words) do to form an intact dupiex. 27. DNA duplex formed from DNA of different species. Tnymidylate is a nucleotide found primarily in acids. 28. Non—WatsomCricic, or of triplex DNA strands. pairing; aliows formation -.E . i . .3 K is g i7 raw '-;~ "set Li Self-Test 107 [:0er i7. Uracil is a nitrogenous base found predominantly in acids. 2. Hydrogen peroxide, a byproduct of aerobic metabolism, is. AACCAATTGG'I‘T is a common cause of these in DNA. T’FGGTT AACC AA 3. Carries genetic information from DNA to the ribosomes. 19' Th e increase in UV 1i ght absorption when doubts 4. Contains a purine attached to a phosphorylated ribose; stranded DN A is denatured is referred to as the the base was first isolated from bird manure. effect. 6' Dehydrated: cornpact form 0f DNA 20. A major pyrimidine; has an amino group at O4. 9‘ A major pyrumdme; has a methyl group at 0'5' 21. Uracil attached through N—l to ribose. 11. Describes mRNA that is translated into more than one 22. Purine or pyrimidine base c o val ently bound to furanose Fromm . _ _ through an Nlegzycosieic bond. 12. Determined for a solution of DNA by measuring UV light absorption as a function of temperature. (2 words) Do You Know the Facts? I. Draw the following structures. _ (a) Adenylate (h) Deoxyguanylate (c) Deoxycytidylate (d) Uridylate (e) Deoxythyrrudylate (f) Hypoxanthine 2 Compounds that contain a nitrogenous base, a pentose sugar, and a phosphate group are called (a) WWW. ’lWo purines found in DNA are (b) M and (c) _..._._ .In DNA, the base pair {(1) __ - is held together by three hydrogen bonds; the base pair (e) _... — W has only two such bonds. In a solution of DNA, the purine and pyrimidine bases stack like coins because of their (1’) hydro Wm nature. When the solution is heated, the base stacking is disrupted, and the optical ab~ sorbance (at 260 rim) of the solution increases; this is called the (g) ' Indicate Whether the following statements about DNA are true or false. (3.) A-form and Bwform DNA are rightmhanded helices, but Z—form DNA is a left—handed helix found only _ in smgle-stranded DNA. b“. (b) Palindroruic sequences can potentially form cruciform structures. (c) The dideoxy method of sequencing DNA can be used on B—DNA, but not on Z—DNA. m—w. (d) Deoxy‘ribose is bound to the nitrogenous base at 01’. 3 (e) A pyrimidine in one strand of DNA always hydrogen bonds with a purine in the opposite strand. 45—”. (f) GEO pairs share three glycosidic bonds. ~—« (g) All the monosaccharide units lack an mOH at 02’. __._ (h) The phosphodiester bonds that link adjacent nucleotides join the 3’ hydroxyl of one nucleotide to the . 5’ hydroxyl of the next. (i) The two strands are aligned as parallei strands. 108 4. 5. 10. . in the DNA sequencing by the Sanger (dideoxy) method: Chapter 8 Nucleotides and Nucleic Acids _._.. (j) The phosphodiester bonds that link adjacent nucleotides are uncharged under physiologicai conditions. ._...._ (k) B—form DNA predominates in aqueous solution; dehydration favors the A—forrn. .m_._ (I) A—i‘orn'i DNA is shorter and has a larger diameter than the B—form DNA. (In) Nucleotide sequence has little or no effect on which form DNA takes. l- .. ADP contains: %- l '5 A. a furanose ring. B. a ketose sugar. C. two phosphoanhydride bonds. D. a B—O-glycosidic bond. E. a pyrimidine base. Which of the following is true for DNA? A. Phosphate groups project toward the middle of the double helix. B. Deoxyribose units are connected by 3',5’—phosphoanhydride bonds. C. The 5' ends of both strands are at the same end of the double helix. D. G50 pairs share three hydrogen bonds. E. The ratio of A+T to G+C is constant for all naturally occurring DNA. A. the template strand of DNA is radioactive. B. enzymes are used to cut the DNA into small pieces, which are then separated by electrophoresis. C. ddT’l‘P is added to each of four reaction mixtures prior to synthesis of complementary strands. D. the role of ddATP is to occasionally terminate synthesis of DNA where dT occurs in the template strands. E. the sequence is read from the top of the gel downward. in living cells, various nucleotides and poiynucleotides: A. serve as structural components of enzyme cofactors. B. alter the equilibria of chemical reactions. 0. carry metabolic energy. D. serve as intracellular signaling molecules. E. all of the above are true. The compound that consists of ribose linked by an N—B-glycosidic ‘nond to N-Q of adenine is: A. a purine nucleotide. B. a pyrimidine nucleotide. C. adenosine. D. AMP. E. deoxyadenosine. The phosphodiester bonds that link adjacent nucleotides in DNA: A. are positively charged. B. join the 3’ hydroxyl of one nucleotide to the 5' hydroxyl of the next. C. always link A with T and C with G. D. are positively charged and always link A with T and C with G. E. are positively charged and join the 3' hydroxyl of one nucleotide to the 5' hydroxyl of the next. Nucleic acid samples have been isolated from three different organisms. The nucleic acids have the following base ratios 0%): 12. 13. 14. I. II. Self-"Fest 109 Which sample (5) are DNA? Which sample would you expect to have the highest Em (melting point)? A. 1 and 3; 3 8. 1,2, and 3;:3 C. 1; 2 D. l and 8; l E. 2 and 3; 2 Two molecuies of double-stranded DNA are the same length (1,000 base pairs), but differ ‘m base composition. Molecule 1 contains 20% A+T; molecule 2 contains 60% A+T. Which molecule has a higher tm (melting point)? How many C residues are there in the 60% AeT DNA molecule? A. 2; 40 B. l; 200 C. 2; 400 D. l; 400 E. 2; 200 What crucial pieces of information from the Hershey-Chase experiment indicated that DNA is the genetic material? Which of “Chargaff’s rules" was the most important clue leading to the model postulated by Watson and Crick? Why? Why does DNA contain thymine rather than uracii? Applying What You Know You wish to determine the relative degree of sequence similarity for DNA from different species of Galapagos finches (referred to as species A, B, and C). You make hybrid duplexes of the DNA between A and 13, A and C, and B and C. You measure the increase in the absorption of UV light with increasing temperature for each of the duplex solutions and obtain the following data: (a) Winch two species have a greater degree of similarity in their DNA sequences? Why? (b) What factors should be taken into account when interpreting these data? How would you use sequence information for a gene in Drosophila to identify and localize a similar gene from a mouse? - Why would reducing the concentration of salts (lowering the ionic strength) in a solution of doubie-stranded DNA lower the melting point of the DNA? 110 Chapter 8 Nucieotides and Nucleic Acids _. Biochemistry on the Internet Nucleic acids are a...
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