Sugars to Nucleotides

1. Sugars to Nucleotides • Last lecture, the role of sugar nucleotides in carbohydrate biosynthesis was described. • Also, the role of ATP in energy metabolism has been emphasized. • Various cofactors have nucleotide character • Later, we see a role for them in cell signaling pathways • Now, we will look primarily at their role in information processing and storage as RNA and DNA

2. The basics of DNA and RNA

3. RNA, a multi-functional molecule • mRNA (messenger RNA) codes for proteins • rRNA (ribosomal RNA) performs peptide bond catalysis in protein synthesis • tRNA (transfer RNA) specify incorporation of amino acids into a protein • additional catalytic and functional RNA molecules (anti-sense, Rnase P, etc.) Although single stranded, RNA can adopt various structures that are important to function

4. Nucleotides have three components • Nitrogenous base • Pentose (ribose or deoxyribose) • Phosphate • The molecule without the phosphate is called a nucleoside

5. The bases are known as pyrimidines and purines

6. Purines and pyrimidines of DNA and RNA

7. Other bases can be found in RNA and DNA • Pseudouridine • Methylcytidine • Methylation of DNA nucleotides (most notably C) is a key aspect of eukaryotic gene expression patterns and adds to the information content of genomic DNA Etc.

8. Although b-furanoses, the sugars differ between RNA and DNA

9. Nucleotides are linked via phosphodiester linkages • Bridges the 5’ hydroxyl group of one sugar and 3’ hydroxyl of the next Phosphate groups are completely ionized at pH 7, thus negatively charged (complexed with metals, etc.)

10. Phosphate groups do not only appear at 3’ and 5’ positions of sugars • 3’, 5’ cAMP is a key intra- and extracellular signal for many biological processes

11. Pyrimidines and Purines have chemical properties that affect structure and function • Planar, or nearly planar • Resonance leads all nucleotide bases to maximum absorption at 260 nm (contrast with 280 nm for protein); Beer’s Law

12. Continued. • Hydrophobic characteristics leads to hydrophobic stacking interactions between bases • Functional groups such as ring nitrogens, carbonyl groups and exocyclic amino groups allow for H-bonding

13. H-bonding leads to complementary base pairing

14. DNA has distinctive, non-random base composition • In all DNA, regardless of species, the number of A’s equals # of T’s, and # G’s = # C’s, such that A + G = T + C • DNA specimens from different tissues of same organism have same base composition • Base composition of DNA can vary wildly among organisms (25% GC vs. 80% GC) • Non-randomness generates signals

15. DNA is a double helix, comprised of anti-parallel strands

16. DNA structure • The hydrophilic backbones of deoxyribose and phosphates are on the outside of the double helix, facing water • The bases are stacked inside the double helix • The glycosidic bonds holding the bases in each basepair are not directly across from one another, hence the sugar-phosphate backbones are not equally spaced yielding a major and minor groove

17. Some proteins bind DNA by recognizing H-bonding patterns by the edges of bases in these grooves

18. DNA has three different forms • B-form DNA: Watson-Crick structure- most stable under physiological conditions; one turn per 3.4 angstroms • A-Form DNA – unclear if a physiological form, only observed in test tube • Z-form DNA – Left-handed helical rotation; Alternating C, G bases can adopt this form in the cell, barely any major groove, minor groove is narrow and deep

20. DNA tertiary structure

21. Enzymes modulate DNA supercoiling • Topoisomerases (gyrase)

22. Nucleic acid structure can be disrupted • Similar to proteins, by heating, or change in pH, one can denature nucleic acid structures • Hydrogen bonds are broken, loss of base-stacking interactions cause strands of DNA double helix to separate • The strands can anneal once temperature or pH is returned to an appropriate temperature • dsDNA and ssDNA have distinct absorbance properties

23. Specific DNA sequences can be synthesized (e.g. primers)

24. Cyclical denaturation and renaturation of DNA is basis of PCR

25. Understanding the significance of DNA sequences provides valuable insight into biology • Reactions terminated by dideoxy NTP’s

26. Era of genome sequencing

27. Sequence data

28. Big Biology

29. Infer Metabolism from Genomes • http://www.genome.jp/kegg/ • Click on KEGG gene universe • Click on PATHWAY • Click on Glycolysis/Gluconeogenesis • Reactants/Products/Enzymes/Pathways

30. Don’t forget proteins are associated with nucleic acids (e.g. histones)

31. Histones affect gene expression

32. RNA has complex structure

33. RNA structure • RNA does not have simple secondary structure such as DNA’s double helix • G:U base pairs are prevalent in RNA in addition to one’s found in DNA • Like proteins, RNA 3-D structure is a complex network of various interactions, most prominently base-stacking

34. Hybridization is the key for microarray or “gene chip” technologies in big biology