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CHAPTER 8 DNA, RNA, and the Flow of Genetic Information Information Transfer in Cells The Central Dogma Polymeric Structure of Nucleic Acids The sequence of the bases constitutes a form of linear information.  Ribose and Deoxyribose Carbon atoms in sugar units are num...

CHAPTER 8 DNA, RNA, and the Flow of Genetic Information Information Transfer in Cells The Central Dogma Polymeric Structure of Nucleic Acids The sequence of the bases constitutes a form of linear information.  Ribose and Deoxyribose Carbon atoms in sugar units are numbered with primes (‘) to distinguish them from carbon atoms in bases Sugar–Phosphate Backbones of Nucleic Acids Purines and Pyrimidines  N-β-Glycosidic Linkages The C-1′ of the sugar is attached to the N-9 of a purine or the N-1 of a pyrimidine by N-β-glycosidic linkage. 10.2 What Are Nucleosides? The base is linked to the sugar via a glycosidic bond The carbon of the glycosidic bond is anomeric Named by adding -idine to the root name of a pyrimidine or -osine to the root name of a purine Conformation can be syn or anti Sugars make nucleosides more water-soluble than free bases 10.2 What Are Nucleosides? Figure 10.10 The common ribonucleosides. Syn and Anti nucleosides Adenosine: A Nucleoside with Physiological Activity Adenosine functions as an autacoid, or local hormone, and neuromodulator. Circulating in the bloodstream, it influences blood vessel dilation, smooth muscle contraction, neurotransmitter release, and fat metabolism. Adenosine is also a sleep regulator. Adenosine rises during wakefulness, promoting eventual sleepiness. Caffeine promotes wakefulness by blocking binding of adenosine to its neuronal receptors. 10.3 What Is the Structure and Chemistry of Nucleotides? Nucleotides are nucleoside phosphates Know the nomenclature "Nucleotide phosphate" is redundant! Most nucleotides are ribonucleotides Nucleotides are polyprotic acids 10.3 What Is the Structure and Chemistry of Nucleotides? Figure 10.11 Structures of the four common ribonucleotides – AMP, GMP, CMP, and UMP. Also shown: 3’-AMP. 10.3 What Is the Structure and Chemistry of Nucleotides? Figure 10.12 The cyclic nucleotide cAMP. 10.3 What Is the Structure and Chemistry of Nucleotides? Figure 10.12 The cyclic nucleotide cGMP Table 4.1 Reproduced from: Biochemistry by T.A. Brown, ISBN: 9781907904288 © Scion 10.3 What Is the Structure and Chemistry of Nucleotides? Figure 10.13 Formation of ADP and ATP by the succesive addition of phosphate groups via phosphoric anhydride linkages. Note that the reaction is a dehydration synthesis. 10.3 What Is the Structure and Chemistry of Nucleotides? Figure 10.13 Formation of ADP and ATP by the succesive addition of phosphate groups via phosphoric anhydride linkages. Note that the reaction is a dehydration synthesis. Nucleoside 5'-Triphosphates Are Carriers of Chemical Energy Nucleoside 5'-triphosphates are indispensable agents in metabolism because their phosphoric anhydride bonds are a source of chemical energy Bases serve as recognition units Cyclic nucleotides are signal molecules and regulators of cellular metabolism and reproduction ATP is central to energy metabolism GTP drives protein synthesis CTP drives lipid synthesis UTP drives carbohydrate metabolism Nucleoside 5'-Triphosphates Are Carriers of Chemical Energy Figure 10.14 Phosphoryl, pyrophosphoryl, and nucleotidyl group transfer, the major biochemical reactions of nucleotides. Phosphoryl group transfer is shown here. Nucleoside 5'-Triphosphates Are Carriers of Chemical Energy Figure 10.14 Phosphoryl, pyrophosphoryl, and nucleotidyl group transfer, the major biochemical reactions of nucleotides. Pyrophosphoryl group transfer is shown here. Nucleoside 5'-Triphosphates Are Carriers of Chemical Energy Figure 10.14 Phosphoryl, pyrophosphoryl, and nucleotidyl group transfer, the major biochemical reactions of nucleotides. Nucleotidyl group transfer is shown here.  The Structure of a Nucleic Acid Chain Can Be Simplified Nucleic acid chains are represented by only the identity of the bases. 10.4 What Are Nucleic Acids? 3',5'-Phosphodiester bridges link nucleotides together to form polynucleotide chains. The 5'-ends of the chains are at the top; the 3'-ends are at the bottom. RNA is shown here. 10.4 What Are Nucleic Acids? 3’,5’-phosphodiester bridges link nucleotides together to form polynucleotide chains. The 5’-ends of the chains are at the top; the 3’-ends are at the bottom. DNA is shown here. 10.5 What Are the Different Classes of Nucleic Acids? DNA - one type, one purpose RNA - 3 (or 4) types, 3 (or 4) purposes ribosomal RNA - the basis of structure and function of ribosomes messenger RNA - carries the message for protein synthesis transfer RNA - carries the amino acids for protein synthesis Others: Small nuclear RNA Small non-coding RNAs  Which statement is true? (1 of 2) a. Each phosphodiester bridge in a nucleic acid backbone has a positive charge. b. Nucleosides consist of a base bonded to a sugar. c. The bases in DNA are adenine (A), thymine (T), guanine (G), and uracil (U). d. Nucleic acids are written in the 3′-to-5′ direction. e. RNA contains the sugar deoxyribose, whereas DNA contains ribose. © Macmillan Learning, 2023  Which statement is true? (2 of 2) a. Each phosphodiester bridge in a nucleic acid backbone has a positive charge. *b. Nucleosides consist of a base bonded to a sugar. c. The bases in DNA are adenine (A), thymine (T), guanine (G), and uracil (U). d. Nucleic acids are written in the 3′-to-5′ direction. e. RNA contains the sugar deoxyribose, whereas DNA contains ribose. © Macmillan Learning, 2023 X-Ray Diffraction Photograph of a Hydrated DNA Fiber Watson–Crick Model of DNA  Structures of the Base Pairs Proposed by Watson and Crick When guanine pairs with cytosine and adenine with thymine, the base pairs have essentially the same shape. Base pairs are held together by weak hydrogen bonds.  If a segment of DNA has the sequence ATCGGCTAAGC, what is the complementary sequence (written in the 3′-to-5′ direction)? (1 of 2) © Macmillan Learning, 2023  If a segment of DNA has the sequence ATCGGCTAAGC, what is the complementary sequence (written in the 3′-to-5′ direction)? (2 of 2) TAGCCGATTCG © Macmillan Learning, 2023  Base Compositions Experimentally Determined for a Variety of Organisms In the 1940s, Erwin Chargaff observed that the A:T and G:C ratios were each nearly 1:1 in a variety of organisms while the A:G ratio varied. TABLE 8.1 Base compositions experimentally determined for a variety of organisms Organism A :T G: C A: G Human being 1.00 1.00 1.56 Salmon 1.02 1.02 1.43 Wheat 1.00 0.97 1.22 Yeast 1.03 1.02 1.67 Escherichia coli 1.09 0.99 1.05 Serratia 0.95 0.86 0.70 marcescens  Stacking of Base Pairs Base stacking contributes to the stability of the double helix because: – double helix formation is facilitated by the hydrophobic effect. – stacked base pairs attract one another through van der Waals forces.  Exercise Biochemist Erwin Chargaff was the first to discover that, in DNA, [A]=[T] and [G]=[C]. These equalities are now known as Chargaff's rule. Using Chargaff's rule, determine the percentages of all of the bases in DNA that is 30% thymine. B-Form, A-Form, and Z-Form DNA  Sugar Pucker Explains Many Structural Differences Between B- Form and A-Form DNA In A-DNA, C-3′ lies out of the plane formed by the other four atoms of the ring (C-3′ endo). – leads to an 11-degree tilting of the base pairs In B-DNA, C-2′ lies out of the plane (C-2′ endo).  Comparison of A-, B-, and Z-DNA TABLE 8.2 Comparison of B-, A-, and Z-DNA B A Z Shape Intermediate Broadest Narrowest Rise per base pair 3.4 Å 2.3 Å 3.8 Å Helix diameter ~20 Å ~26 Å ~18 Å Screw sense Right-handed Right-handed Left-handed Glycosidic bond* anti anti Alternating anti and syn Base pairs per turn of helix 10.4 11 12 Pitch per turn of helix 35.4 Å 25.3 Å 45.6 Å Tilt of base pairs from 1 degree 19 degrees 9 degrees perpendicular to helix axis Sugar puckering C-2′ endo C-3′ endo Mixed *Syn and anti refer to the orientation of the N-glycosidic bond between the base and deoxyribose. In the anti orientation, the base extends away from the deoxyribose. In the syn orientation, the base is above the deoxyribose. Pyrimidines can be in anti orientations only, whereas purines can be anti or syn. The Major and Minor Groove Relaxed and Supercoiled Circular DNA Forms Stem-Loop Structures Identify a stem loop structure. (1 of 2) © Macmillan Learning, 2023 Identify a stem loop structure. (2 of 2) © Macmillan Learning, 2023 Complex Structure of an RNA Molecule The Chemical Differences Between DNA and RNA Have Biological Significance Figure 10.25 Deamination of cytosine forms uracil. The Chemical Differences Between DNA and RNA Have Biological Significance Figure 10.26 The 5-methyl group on thymine labels it as a special kind of uracil. Depurination of DNA by dilute acid -a proton attacks the N7 nitrogen. -the N-glycosidic bond is broken. -the oxocarbenium ion formed is hydroxylated by a water molecule. RNA is less susceptible due to its shorter glycosidic bond.  Hydrolysis of RNA by dilute base - an OH- abstracts a proton from the C’2 hydroxyl group, leaving a charged oxygen. - The charged oxygen attacks the positive phosphorus breaking the phosphodiester bond on the “b” side and forming a cyclic monophosphate derivative. - the cyclic nucleotide is hydrolyzed by water either on the C’2 or C’3 side to for a mixture of 2’- and 3’-monophosphate derivatives. Semiconservative Replication Hypochromism  DNA replication: (1 of 2) a. is called conservative because each new helix retains both of the parental strands. b. only occurs when the temperature is increased above the Tm. c. requires hydrogen bonds to be disrupted. d. is accompanied by a decrease in absorption of 260 nm light. e. results in second-generation daughter molecules that do not contain any of the original parental strands. © Macmillan Learning, 2023  DNA replication: (2 of 2) a. is called conservative because each new helix retains both of the parental strands. b. only occurs when the temperature is increased above the Tm. *c. requires hydrogen bonds to be disrupted. d. is accompanied by a decrease in absorption of 260 nm light. e. results in second-generation daughter molecules that do not contain any of the original parental strands. © Macmillan Learning, 2023  Section 8.4 DNA Is Replicated by Polymerases That Take Instructions from Templates DNA polymerase catalyzes phosphodiester-bridge formation in a step-by-step manner (DNA)n + dNTP ⇌ (DNA)n+1 + PPi where dNTP is any deoxyribonucleotide and PPi is a pyrophosphate ion Key Characteristics of DNA Synthesis key characteristics include: – the reaction requires four deoxynucleoside 5′-triphosphates and Mg2+. – the new DNA strand is assembled on a preexisting DNA template (the template strand) – DNA polymerases require a primer to begin synthesis – chain elongation proceeds in the 5′-to-3′ direction – many DNA polymerases have nuclease activity to remove mismatched nucleotides DNA Polymerases Catalyze Strand Elongation  The Genes of Some Viruses Are Made of RNA Some viruses have RNA genomes that are replicated by RNA-directed RNA polymerases. – example = tobacco mosaic virus retroviruses = viruses with single-stranded RNA genomes that are converted to DNA double helices by reverse transcriptase – example = human immunodeficiency virus 1 (HIV-1) Flow of Information from RNA to DNA in Retroviruses  Section 8.5 Gene Expression Is the Transformation of DNA Information into Functional Molecules DNA is expressed in two steps: – Step 1: an RNA copy (messenger RNA, mRNA) is made that encodes directions for protein synthesis – Step 2: information in mRNA is translated to synthesize functional proteins  Several Kinds of RNA Play Key Roles in Gene Expression major kinds of RNA that are involved in gene expression: – messenger RNA (mRNA) = template for protein synthesis – transfer RNA (tRNA) = carries amino acids in an activated form to the ribosome for peptide-bond formation – ribosomal RNA (rRNA) = major component of ribosomes that serves as the actual catalyst for protein synthesis  RNA Molecules in E. coli TABLE 8.3 RNA molecules in E. coli Sedimentation Number of Type Relative amount (%) coefficient (S) Mass (kDa) nucleotides Ribosomal RNA 23 1.2 × 103 3700 (rRNA) 80 16 0.55 × 103 1700 5 3.6 × 101 120 Transfer RNA (tRNA) 15 4 2.5 × 101 75 Messenger RNA (mRNA) 5 Heterogeneous  All Cellular RNA Is Synthesized by RNA Polymerases transcription = synthesis of RNA from a DNA template RNA polymerase catalyzes the initiation and elongation of RNA chains (RNA)n residues + ribonucleoside triphosphate ⇌ (RNA)n+1 residues + PPi  RNA Polymerase RNA polymerase doesn't require a primer.  Key Requirements of RNA Polymerase RNA polymerase requires: – a double- or single-stranded DNA template. – four ribonucleoside triphosphates (ATP, GTP, UTP, and CTP). – a divalent metal ion (Mg2+ or Mn2+). RNA and DNA Synthesis Similarities The direction of synthesis is in the 5′-to-3′ direction. mechanism of elongation = 3′–OH at the terminus of the growing chain makes a nucleophilic attack on the innermost phosphoryl group of the incoming nucleoside triphosphate Synthesis is driven by pyrophosphate hydrolysis. RNA Polymerase Catalyzes the Strand-Elongation Reaction RNA Polymerases Take Instructions from DNA Templates Early evidence found that TABLE 8.4 Base composition (percentage) of RNA synthesized the base composition of from a viral DNA template the newly synthesized DNA template RNA is the complement (plus, or coding, RNA of the DNA template strand of ϕX174) product strand. A 25 U 25 T 33 A 32 G 24 C 23 C 18 G 20 Complementarity Between mRNA and DNA If a template strand of DNA has the sequence 3′–TCAAGGCGA–5′, what is the corresponding mRNA sequence (written in the 5′-to-3′ direction)? (1 of 2) © Macmillan Learning, 2023 If a template strand of DNA has the sequence 3′–TCAAGGCGA–5′, what is the corresponding mRNA sequence (written in the 5′-to-3′ direction)? (2 of 2) 5′–AGUUCCGCU–3′ © Macmillan Learning, 2023 Transcription Begins Near Promoter Sites and Ends at Terminator Sites promoter sites = regions along DNA templates that specifically bind RNA polymerase and determine where transcription begins – typically vary from an idealized single sequence, or consensus sequence, by only 1–2 residues – prokaryote examples: Pribnow box, –35 region – eukaryote examples: TATA box (Hogness box), CAAT box Promoter Sites for Transcription in Prokaryotes and Eukaryotes  Transcription Termination Prokaryote termination occurs when RNA polymerase synthesizes a terminator sequence or by the action of the protein rho. Terminator sequence is a stem-loop structure followed by a sequence of U residues. – Structure forms by base-pairing of self-complementary sequences that are rich in G and C. Less is known about eukaryote termination. A Stem-Loop Structure at the 3′ End of an E. coli mRNA Transcript  Modification of Eukaryotic mRNA in eukaryotes, mRNA is modified: – a "cap" structure (guanosine nucleotide attached to mRNA with a 5′-5′ triphosphate linkage) is attached to the 5′ end – a sequence of adenylates (a poly(A) tail) is added to the 3′ end  Transfer RNAs Are the Adaptor Molecules in Protein Synthesis Transfer RNAs (tRNAs): – bring amino acids to the mRNA. – contain an amino-acid attachment site and a template recognition site. – have several regions of base-paired segments in multiple stem loons. – are called aminoacyl-tRNAs when an amino acid is attached by an aminoacyl-tRNA synthetase. codon = three coding bases on the mRNA template anticodon = three complementary bases on the tRNA General Structure of an Aminoacyl- tRNA  Attachment of an Amino Acid to a tRNA Molecule The amino acid is attached to 3′–OH group of the ribose at the 3′ end of the tRNA molecule. CCA terminus = region at the 3′ end tRNA sequence that contains two cytidylates followed by an adenylate Section 8.6 Amino Acids Are Encoded by Groups of Three Bases Starting from a Fixed Point genetic code = the relation between the sequence of bases in DNA and the sequence of amino acids in proteins Features of the genetic code: – three nucleotides encode an amino acid – nonoverlapping – has no punctuation – has directionality – is degenerate (most amino acids are encoded by more than one codon)  The Genetic Code TABLE 8.5 The genetic code Sixty-one First Second position Third position triplets specify position (5' end) U Phe C Ser A Tyr G Cys (3' end) U amino acids. U Phe Ser Tyr Cys C Leu Ser Stop Stop A Three triplets Leu Leu Ser Stop Pro His Trp Arg G U are stop C Leu Leu Pro Pro His Gln Arg Arg C A codons that Leu lle Pro Thr Gln Asn Arg Ser G U designate A lle Thr Asn Ser C termination of lle Thr Lys Arg A Met Thr Lys Arg G translation. G Val Val Ala Ala Asp Asp Gly Gly U C Val Ala Glu Gly A Val Ala Glu Gly G Note: This table identifies the amino acid encoded by each triplet. For example, the codon 5'-AUG-3' on mRNA specifies methionine, whereas CAU specifies histidine. UAA, UAG, and UGA are termination signals. AUG is part of the initiation signal, in addition to coding for internal methionine residues.  For the codon AAA, click on the amino acid. (1 of 2) TABLE 8.5 The genetic code First Second position Third position position (5' end) U C A G (3' end) Phe Ser Tyr Cys U U Phe Ser Tyr Cys C Leu Ser Stop Stop A Leu Ser Stop Trp G Leu Pro His Arg U C Leu Pro His Arg C Leu Pro Gln Arg A Leu Pro Gln Arg G lle Thr Asn Ser U A lle Thr Asn Ser C lle Thr Lys Arg A Met Thr Lys Arg G Val Ala Asp Gly U G Val Ala Asp Gly C Val Ala Glu Gly A Val Ala Glu Gly G Note: This table identifies the amino acid encoded by each triplet. For example, the codon 5'-AUG-3' on mRNA specifies methionine, whereas CAU specifies histidine. UAA, UAG, and UGA are termination signals. AUG is part of the initiation signal, in addition to coding for internal methionine residues.  For the codon AAA, click on the amino acid. (2 of 2) TABLE 8.5 The genetic code First Second position Third position position (5' end) U C A G (3' end) Phe Ser Tyr Cys U U Phe Ser Tyr Cys C Leu Ser Stop Stop A Leu Ser Stop Trp G Leu Pro His Arg U C Leu Pro His Arg C Leu Pro Gln Arg A Leu Pro Gln Arg G lle Thr Asn Ser U A lle Thr Asn Ser C lle Thr Lys Arg A Met Thr Lys Arg G Val Ala Asp Gly U G Val Ala Asp Gly C Val Ala Glu Gly A Val Ala Glu Gly G Note: This table identifies the amino acid encoded by each triplet. For example, the codon 5'-AUG-3' on mRNA specifies methionine, whereas CAU specifies histidine. UAA, UAG, and UGA are termination signals. AUG is part of the initiation signal, in addition to coding for internal methionine residues.  The Genetic Code Is Highly Degenerate synonyms = codons that specify the same amino acids – most differ only in the last base of the triplet biological significance of degeneracy: – decreases probability of mutating to chain termination – minimizes the deleterious effects of mutations  Codon Bias codon bias = nonrandom use of synonymous codons in different organisms – may help regulate translation  Messenger RNA Contains Start and Stop Signals for Protein Synthesis ribosomes = large molecular complexes assembled from protein and ribosomal RNA mRNA is translated into proteins on ribosomes. Stop codons are read by proteins called release factors rather than tRNA molecules. The Start Signal for Protein Synthesis In prokaryotes: – an initiator tRNA carries a modified amino acid, formylmethionine (fMet), and recognized AUG. – the initiating AUG codon is preceded by the purine-rich Shine–Dalgarno sequence which base pairs with a complementary sequence in ribosomal RNA. In eukaryotes, the AUG nearest the 5 end is the initiator codon. reading frame = order of the three nonoverlapping nucleotides – established by the location of the initiator codon Initiation of Protein Synthesis The Genetic Code Is Nearly Universal Most organisms use the same genetic code because there is strong selection against deleterious mutations. Some genomes are translated by different code – example: in ciliated protozoa, codons that are stop codons in most organisms encode amino acids – example: mitochondria also use variations in the genetic code because mitochondrial DNA encodes a distinct set of tRNAs that recognize alternative codons Distinctive Codons of Human Mitochondria TABLE 8.6 Distinctive codons of human mitochondria Codo Standard Mitochondrial n Code code UGA Stop Trp UGG Trp Trp AUA lle Met AUG Met Met AGA Arg Stop AGG Arg Stop  Section 8.7 Most Eukaryotic Genes Are Mosaics of Introns and Exons eukaryotic genes are discontinuous – exons = coding regions – introns = noncoding regions The average human gene has 8 introns, while some have more than 100. Intron size ranges from 50 to 10,000 nucleotides. RNA Processing Generates Mature RNA Eukaryotic pre-messenger RNA (pre-mRNA) contains exons and introns. Following modifications, introns are spliced out and coding sequences are linked at the 3' end. spliceosomes = assemblies of proteins and small nuclear RNA molecules (snRNAs) that carry out splicing  The Spliceosome Recognizes Specific Sequences Within the Intron That Specify the Splice Sites Introns almost always begin with a GU and end with an AG that is preceded by a pyrimidine-rich tract.  Most genes in eukaryotes: (1 of 2) a. are continuous. b. are transcribed and spliced to generate a primary transcript, which is then modified by cap and poly(A) addition. c. are randomly spliced to generate multiple proteins. d. contain introns that may or may not encode for protein. e. contain exons ordered in the same sequence in mRNA as in DNA. © Macmillan Learning, 2023  Most genes in eukaryotes: (2 of 2) a. are continuous. b. are transcribed and spliced to generate a primary transcript, which is then modified by cap and poly(A) addition. c. are randomly spliced to generate multiple proteins. d. contain introns that may or may not encode for protein. *e. contain exons ordered in the same sequence in mRNA as in DNA. © Macmillan Learning, 2023 Many Exons Encode Protein Domains Many exons encode discrete structural elements, binding sites, and catalytic sites. exon shuffling = process by which new proteins arise in evolution by the rearrangement of exons alternative splicing = process allowing the generation of multiple proteins from a primary transcript  The Tissue Plasminogen Activator (TPA) Gene Was Generated by Exon Shuffling  Alternative Splicing Alternative splicing provides a means of forming a set of proteins that are variants of a basic motif without requiring a separate gene for each protein. Remember to complete your exit poll tonight! © Macmillan Learning, 2023