BCH 5045 Graduate Survey of Biochemistryhort.ifas.ufl.edu/faculty/guy/bch5045/Lecture Files/Lecture...

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BCH 5045

Graduate Survey of Biochemistry

Instructor: Charles Guy Producer: Ron Thomas Director: Glen Graham

Lecture 16

Slide sets available at: http://hort.ifas.ufl.edu/teach/guyweb/bch5045/index.html

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• LEHNINGER • PRINCIPLES OF BIOCHEMISTRY

• Fifth Edition

David L. Nelson and Michael M. Cox

© 2008 W. H. Freeman and Company

CHAPTER 8

Nucleotides and Nucleic Acids



Presenter

Presentation Notes

FIGURE 8-14 Complementarity of strands in the DNA double helix. The complementary antiparallel strands of DNA follow the pairing rules proposed by Watson and Crick. The base-paired antiparallel strands differ in base composition: the left strand has the composition A3 T2 G1 C3; the right, A2 T3 G3 C1. They also differ in sequence when each chain is read in the 5′→3′ direction. Note the base equivalences: A = T and G = C in the duplex. FIGURE 8-1b Structure of nucleotides. (b) The parent compounds of the pyrimidine and purine bases of nucleotides and nucleic acids, showing the numbering conventions. FIGURE 8-2 Major purine and pyrimidine bases of nucleic acids. Some of the common names of these bases reflect the circumstances of their discovery. Guanine, for example, was first isolated from guano (bird manure), and thymine was first isolated from thymus tissue.



The building blocks of nucleic acids!

Presenter

Presentation Notes

FIGURE 8-1a Structure of nucleotides. (a) General structure showing the numbering convention for the pentose ring. This is a ribonucleotide. In deoxyribonucleotides the —OH group on the 2′ carbon (in red) is replaced with —H.



What is this monosaccharide and what does it have to do with DNA?

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Presentation Notes

FIGURE 8-3a Conformations of ribose. (a) In solution, the straight-chain (aldehyde) and ring (β-furanose) forms of free ribose are in equilibrium. RNA contains only the ring form, β-D-ribofuranose. Deoxyribose undergoes a similar interconversion in solution, but in DNA exists solely as β-2′-deoxy-D-ribofuranose. FIGURE 8-3b Conformations of ribose. (b) Ribofuranose rings in nucleotides can exist in four different puckered conformations. In all cases, four of the five atoms are in a single plane. The fifth atom (C-2′ or C-3′) is on either the same (endo) or the opposite (exo) side of the plane relative to the C-5′ atom.



The building blocks of DNA and RNA. What is the major difference between DNA and RNA and what biochemical significance does it have?

The estimated half-life of the 3’ P---O—C part of the phosphodiester bond to hydrolytic cleavage by water in DNA at 25°C is in the neighborhood of 31 million years while the same bond in RNA has a half-life of about 1 year. See commentary on next slide.

Presenter

Presentation Notes

FIGURE 8-4a Deoxyribonucleotides and ribonucleotides of nucleic acids. All nucleotides are shown in their free form at pH 7.0. The nucleotide units of DNA (a) are usually symbolized as A, G, T, and C, sometimes as dA, dG, dT, and dC; those of RNA (b) as A, G, U, and C. In their free form the deoxyribonucleotides are commonly abbreviated dAMP, dGMP, dTMP, and dCMP; the ribonucleotides, AMP, GMP, UMP, and CMP. For each nucleotide, the more common name is followed by the complete name in parentheses. All abbreviations assume that the phosphate group is at the 5′ position. The nucleoside portion of each molecule is shaded in pink. In this and the following illustrations, the ring carbons are not shown. FIGURE 8-4b Deoxyribonucleotides and ribonucleotides of nucleic acids. All nucleotides are shown in their free form at pH 7.0. The nucleotide units of DNA (a) are usually symbolized as A, G, T, and C, sometimes as dA, dG, dT, and dC; those of RNA (b) as A, G, U, and C. In their free form the deoxyribonucleotides are commonly abbreviated dAMP, dGMP, dTMP, and dCMP; the ribonucleotides, AMP, GMP, UMP, and CMP. For each nucleotide, the more common name is followed by the complete name in parentheses. All abbreviations assume that the phosphate group is at the 5′ position. The nucleoside portion of each molecule is shaded in pink. In this and the following illustrations, the ring carbons are not shown.



The Genomes of Neandertals and Modern Humans As indicated on the previous slide, DNA is a pretty stable molecule for good reason, and this is largely why it is possible to obtain DNA or genomic sequence information from something or someone who lived 10s of thousands of years ago. All that is needed is the right environmental and microbiological conditions for antediluvian DNA, even highly contaminated with bacterial DNA, to be sequencible by modern methods and technologies. In 2010 Reich, Pääbo and colleagues reported on the analysis of nuclear DNA sequences extracted from a finger bone found in Denisova Cave in southern Siberia. They conclude that their sequencing results suggest an unexpected complex history of migration and colonization of Europe and Asia after modern humans presumably moved out of Africa 50,000–60,000 years ago (1). Previously, genetic data and the fossil record favored a so called “replacement model” with respect to Neandertals (or Neanderthal either way is correct) in which modern humans derive their greatest genetic ancestry to a breeding population with an origin in Africa about 200,000 years ago (2, 3, 4, 5), but not from Neandertals. The new nuclear genome sequence from the Siberian cave (1) and another sequence for Homo neanderthalensi (6) suggest that the out-of-Africa replacement hypothesis may need revision.

This slide not in the lecture video



It appears that the Denisovan sequence and previously determined Neandertal sequences are indeed related, but are not the same. Instead the Denisovan sequence is more closely similar to modern humans than has been found in other Neandertal sequences. Now the interesting part is that the Denisovan sequences cluster slightly better with present-day European or East Asian genomes than with African genomes, which is consistent with the view of some gene flow from Neanderthals to the progenitors of the ancestors of modern-day Eurasians (6). Unexpectedly the Denisovan Cave sequence seems to show a higher genetic relatedness with present-day island Melanesians. Overall, from the genome sequences it appears that there could have been limited gene flow from Neandertals to modern humans at two different times in different parts of the world, in Eurasia (early) and in Oceania (later). Still Svante Pääbo of the Max Planck Institute for Evolutionary Anthropology in Germany notes that the Neandertal and modern human genomes share 99.5-99.9% nucleotide sequences identity.




The Green et al. report (6) suggests that between 1 to 5% of non-African individuals’ genomes has an actual Neandertal origin, while Reich et al. results (1) suggest that 4 to 6% of the genomes of Melanesians are derived from the sequence from the archaic hominin population finger bone from the Denisova Cave has become known as the Denisovans. Yet, a more recent work (7) on Neandertal genome sequences has indicated an admixture between Neandertals and the expanding population of modern humans, H. sapiens who left Africa between 80,000 and 50,000 years ago to colonize the rest of the world. There is evidence for a significant presence (9% overall) of a Neandertal-derived X chromosome segment in all contemporary human populations outside Africa. Similarly, the analysis of 6092 X-chromosomes from populations that inhabit all continents seems to further support the assertions that a “mosaic of lineages” formed at different times and of different geographic locations contributes to the genetic constitution of modern humans. The analyses indicate an early admixture between an expanding African migrant population and Neandertals prior to or early in the out-of-Africa expansion that led to the successful colonization of the world by modern humans.




The most recent Neandertal site dates to about 28,000 years ago. What happened to them remains unknown, but is the subject of much speculation. 1. Reich, D. et al. Nature 468, 1053–1060 (2010). 2. Cann, R., Stoneking, M. & Wilson, A. Nature 325, 31–36 (1987). 3. Ramachandran, S. et al. Proc. Natl Acad. Sci. USA 102, 15942–15947 (2005). 4. Underhill, P. et al. Ann. Hum. Genet. 65, 43–62 (2001). 5. Klein, R. The Human Career 3rd edn (Univ. Chicago Press, 2009). 6. Green, R. E. et al. Science 328, 710–722 (2010). 7. Yotova, V. et al. Mol Biol Evol. Jul;28(7):1957-62. Epub 2011 Jan 25.




Hold on, there are additional purine and pyrimidine bases that can be found in DNA and RNA that we typically don’t hear much about. Of those shown, which nucleic acid would you expect them to be found?

Presenter

Presentation Notes

FIGURE 8-5a Some minor purine and pyrimidine bases, shown as the nucleosides. (a) Minor bases of DNA. 5-Methylcytidine occurs in the DNA of animals and higher plants, N6-methyladenosine in bacterial DNA, and 5-hydroxymethylcytidine in the DNA of bacteria infected with certain bacteriophages. FIGURE 8-5b Some minor purine and pyrimidine bases, shown as the nucleosides. (b) Some minor bases of tRNAs. Inosine contains the base hypoxanthine. Note that pseudouridine, like uridine, contains uracil; they are distinct in the point of attachment to the riboseﾑin uridine, uracil is attached through N-1, the usual attachment point for pyrimidines; in pseudouridine, through C-5.



Products of RNA hydrolysis. Alkaline conditions promote RNA hydrolysis.

Presenter

Presentation Notes

FIGURE 8-6 Some adenosine monophosphates. Adenosine 2′-monophosphate, 3′-monophosphate, and 2′,3′-cyclic monophosphate are formed by enzymatic and alkaline hydrolysis of RNA.



Tautomers of uracil. Other bases found in nucleic acids have tautomeric forms also. Tautomerization is the interconversion of keto group to alcohol.

Presenter

Presentation Notes

FIGURE 8-9 Tautomeric forms of uracil. The lactam form predominates at pH 7.0; the other forms become more prominent as pH decreases. The other free pyrimidines and the free purines also have tautomeric forms, but they are more rarely encountered.



Nucleic acids have a beginning and an end. How can we tell which is which?

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Presentation Notes

FIGURE 8-7 Phosphodiester linkages in the covalent backbone of DNA and RNA. The phosphodiester bonds (one of which is shaded in the DNA) link successive nucleotide units. The backbone of alternating pentose and phosphate groups in both types of nucleic acid is highly polar. The 5′ end of the macromolecule lacks a nucleotide at the 5′ position, and the 3′ end lacks a nucleotide at the 3′ position.



Nucleic acids have absorbance peaks near 260 nm. This physical property of nucleic acids, especially DNA, has everyday implications and consequences.

Presenter

Presentation Notes

FIGURE 8-10 Absorption spectra of the common nucleotides. The spectra are shown as the variation in molar extinction coefficient with wavelength. The molar extinction coefficients at 260 nm and pH 7.0 (ε260) are listed in the table. The spectra of corresponding ribonucleotides and deoxyribonucleotides, as well as the nucleosides, are essentially identical. For mixtures of nucleotides, a wavelength of 260 nm (dashed vertical line) is used for absorption measurements.



What type of weak bonding interaction between the two strands of DNA shown serve to provide stability to the double helical structure of DNA?

Presenter

Presentation Notes

FIGURE 8-11 Hydrogen-bonding patterns in the base pairs defined by Watson and Crick. Here as elsewhere, hydrogen bonds are represented by three blue lines.



http://www.youtube.com/watch?v=wpoCbd3bb68

http://www.youtube.com/watch?v=wpoCbd3bb68�



Presenter

Presentation Notes

FIGURE 8-13 Watson-Crick model for the structure of DNA. The original model proposed by Watson and Crick had 10 base pairs, or 34 Å (3.4 nm), per turn of the helix; subsequent measurements revealed 10.5 base pairs, or 36 Å (3.6 nm), per turn. (a) Schematic representation, showing dimensions of the helix. (b) Stick representation showing the backbone and stacking of the bases. (c) Space-filling model.

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