1. NUCLEIC ACIDS

2. OBJECTIVES  Identify/ recognize nucleic acid  Components in nucleic acid – monosaccharide, nucleobases, phosphoric acid  Differentiate - between 2 types of nucleic acids, DNA and RNA - between nucleotide and nucleoside - between nucleotide and nucleoside  Definition – nucleotide, nucleoside, DNA and RNA

3. Nucleic Acids  Nucleic acid: a biopolymer containing three types of monomer units –a nitrogenous base (nucleobases), either purine or pyrimidine –a monosaccharide (aldopentose), either D-ribose or 2- deoxy-D-ribose –phosphoric acid/phospharyl group  Two types - RNA (Ribonucleic Acid) - DNA (Deoxyribonucleic Acid) - DNA (Deoxyribonucleic Acid)

4. Nucleobases  Heterocylic compounds containing C, H, N, and O  Purine and pyrimidine methyl Nonpolar (C)(T)(U) (A)(G)

5.  2 type of aldopentoses found - Ribose (RNA) - 2-deoxyribose (DNA)  Deoxyribose, derivative of ribose – lacks an oxygen atom at C2 Monosaccharide/sugar Polar Only  anomer present in nucleic acid

6. Nucleosides Lack phosphate group Pyrimidine Purine  Nucleoside:  Covalent linkage forms between

7. Nucleotides  Nucleotide: a nucleoside in which a molecule of phosphoric acid/phosphoryl group is esterified with an -OH of the monosaccharide, at the 5’-OH  As constituents of cofactors, Coenzyme A (CoA), flavin adenine dinucleotide (FAD) & nicotinamide adenine dinucleotides (NAD) Nucleobase, aldopentose sugar and phosphoryl group Phosphoric acid - polar

8. 5’ = attach to C5 of pentose SUGAR?

9. NOMENCLATURE of Nucleotide Based on the nucleoside, plus the phosphate group

10. Nucleotide Sequence  Gene:  The nucleotide sequence is depending on the bases (nucleobases) present

11. Nucleic Acid: DNARNA 1. Bases = 2. Aldopentose = 3. Phosphoryl group 1. Bases = 2. Aldopentose = 3. Phosphoryl group Biopolymer, nucleotide as monomer Naming of nucleotide: if Base adenine  Deoxyadenosine 5’ monophosphate Naming of nucleotide: if Base adenine  Adenosine 5’monophosphate Nucleoside

12. Nucleic Acid - DNA and RNA  DNA stands for deoxyribonucleic acid. It is the genetic code molecule for most organisms.  RNA stands for ribonucleic acid. RNA molecules are involved in converting the genetic information in DNA into proteins. In retroviruses, RNA is the genetic material. NUCLEIC ACIDS ARE POLYMERS OF NUCLEOTIDES

13. DNA  DNA and RNA are polymers whose monomer units are nucleotides = polynucleotides  Deoxyribonucleic acids, DNA: a biopolymer that consists of a backbone of alternating units of 2- deoxy-D-ribose and phosphoryl group –the 3’-OH of one nucleotide is joined to the 5’ P of the next nucleotide by a phosphodiester bond Polynucleotide = DNA and RNA 3’ 5’ -phosphodiester bond Hydrolysis – break bond Condensation – form bond

14. DNA structure  Levels of structure –1° structure: –2° structure: = double helix structure –3° structure: –4° structure:

15. DNA - 1° Structure  Primary Structure: the sequence of bases along the pentose- phosphodiester backbone of a DNA molecule –base sequence is read from the 5’ end to the 3’ end –System of notation single letter (A,G,C and T) 5’ – G G C A T T G C G C - 3’ On the right 3’ 5’ -phosphodiester bond Pg 237, Campbell and Farrel. READ!

16. Segment of DNA Chain 5’-end 3’-end guanine thymine cytosine 3’-5’ link 5’ end – phosphate group is free 3’end – 3’ OH in deoxyribose is free

17. DNA - 2° Structure  Secondary structure: the ordered arrangement of nucleic acid strands  Double helix: a type of 2° structure of DNA molecules in which two antiparallel polynucleotide strands are coiled in a right-handed manner about the same axis The chains run antiparallel and are held together by hydrogen bonding between complementary base pairs: A=T, G=C.The chains run antiparallel and are held together by hydrogen bonding between complementary base pairs: A=T, G=C. DNA double helix

18. DNA structural elements OHP  2 right-handed, helical, polynucleotide chains, coiled around a common axis to form a double helix  2 characteristic: Major groove and minor groove – binding site for drug or polypeptide  2 strands run in opposite direction (antiparallel)-3’,5’-phosphodiester bridges run in opposite direction  1 base (purine) from single strand link to 1 base (pyrimidine) from other stand (complimentary)  Bases are perpendicular to helix axis  Polarity and non-polarity regions  Aqueous environment – polar, charged, covalent backbone deoxyribose and phosphate groups outside of the helix  Hydrophobic purine and pyrimidine bases avoid water by turning towards the inside of the structure

19. T-A Base Pairing  Base pairing is complementary: A=T, G  C  A major factor stabilizing the double helix is base pairing by hydrogen bonding between T-A and between C-G  T-A base pair comprised of 2 hydrogen bonds Complementary base pairing

20. G-C Base Pair  G-C base pair comprised of 3 hydrogen bonds

21. Forms of DNA  B-DNA –considered the –a right-handed helix, inside diameter 11Å –10 base pairs per turn (34Å) of the helix  A-DNA –a right-handed helix, but thicker than B-DNA –11 base pairs per turn of the helix  Z-DNA a left-handed double helixa left-handed double helix may play a role in gene expressionmay play a role in gene expression Z-DNA occurs in nature, usually consists of alternating purine-pyrimidine bases Z-DNA occurs in nature, usually consists of alternating purine-pyrimidine bases Methylated cytosine found also in Z-DNA Methylated cytosine found also in Z-DNA

22. Structural features of A-, B-, and Z- DNA TypeA-DNAB-DNAZ-DNA Helical sensesright handedright handedleft handed Diameter (Å)~26~20~18 Base pairs/turn111012 Major groovenarrow/deepwide/deepFlat Minor groovewide/shallownarrow/deepnarrow/deep Pg 294, Concepts in Biochemistry. 3/e 2006 John Wiley & Sons 20 Å

23. DNA - 3° Structure  Tertiary structure: the three-dimensional arrangement of all atoms of a nucleic acid; commonly referred to as supercoiling  Supercoiling- Further coiling and twisting of DNA helix.

24. DNA  DNA can forms tertiary structure by twist into complex arrangement – supercoil  Circular DNA:  Can be found in  Circular twisted into supercoiled DNA - 3° Structure  Supercoil - results of extra twisting in the linear duplex form

25. DNA  Circular DNA: In microorganisms (bacteriophages, bacteria)  Circular twisted into supercoiled DNA - 3° Structure  In eukaryotes, the 3° structure involves histone (protein)- Chromatin: DNA molecules wound around particles of histones in a beadlike structure

26. PROPERTIES OF SUPERCOIL  Supercoiled is  Compact  Play a regulatory role in DNA replication

27. Bacteriophage : DNA – threadlike structure

28. Super DNA Coiled Topology  Double helix can be considered to a 2-stranded, right handed coiled rope  Can undergo positive/negative supercoiling clockwise Counterclockwise

29. DNA - 4° Structure  Four stranded form of DNA (quadruplex DNA)  Role in regulating and stabilizing telomeres and in regulation of gene expression  Small molecules such as porphyrins and anthraquinones present, to stabilize the structure G-quadruplex

30. NUCLEIC ACIDS (2)

31. OBJECTIVES  Denaturation of DNA  Identify/ recognize RNA  Differentiate between DNA and RNA  Differentiate between mRNA, tRNA, rRNA

32. DNA  Can be disrupted by heat, acids, bases or organic solvents (double helix denatured = unwinding of the DNA double helix)  In nature, the unwinding of the DNA double helix is the important step in DNA replication Involves the nitrogenous bases

33. Denaturation of DNA  Denaturation: disruption of 2° structure –most commonly by heat denaturation (melting- the heat denaturation of DNA) –as strands separate, absorbance at 260 nm increases –increase is called hyperchromicity- the wavelength of absorption does not change but the amount of light absorbed increases –midpoint of transition (melting) curve = T m –the higher the % G-C, the higher the T m –Renaturation/annealing is possible on slow cooling  Principal use in PCR At the nitrogenous bases

34. Denaturation of DNA  Double helix unwinds when DNA is denatured  Can be re-formed with slow cooling and annealing

35. RNA  consist of long, unbranched chains of nucleotides joined by phosphodiester bonds between the 3’-OH of one pentose and the 5’-P of the next nucleotide  the pentose unit is  -D-ribose (it is 2-deoxy-D- ribose in DNA)- the extra OH present in RNA makes this nucleotide more susceptible to hydrolysis than DNA.  the pyrimidine bases are uracil and cytosine (they are thymine and cytosine in DNA)  RNA is single stranded (DNA is double stranded) The bases sequences of all types of RNA are determined by that of DNA

36. RNA  RNA molecules are classified according to their structure and function

37. RNA structure  Levels of structure –1° structure: the order of bases on the polynucleotide sequence; complementary to the DNA template –2° structure: no specific 2° arrangements, but RNA is not completely lacking of regular structure –3° structure:interaction between DNA and proteins

38. RNA- 1° Structure  Polymer of nucleotide  Involves single polynucleotide strand

39. RNA- 2° Structure  Loop back onto themselves to fold into conformation containing several different structural elements: 1-hairpin turns 2-right-handed double helixes 3-internal loops All classes of RNA synthesized as single- stranded molecules 3’OH 5’P

40.  Hairpin turn - Loops in the single chain  Right-handed double helixes - result of intrastrand folding - Trigger by hairpin turn - Antiparallel & stabilized in the same direction as in DNA - Hold by H bond & stacking interaction  Internal loops - Common in RNA - Structural features that disrupt the formation of continuous double helix regions

41. tRNA  Transfer RNA, tRNA: –the smallest kind of the three RNAs –a single-stranded polynucleotide chain between 73-94 nucleotide residues –carries an amino acid at its 3’ end –intramolecular hydrogen bonding occurs in tRNA Cloverleaf structure Function: Involves in synthesis of polypeptide, to carry amino acid to site of protein synthesis

42. tRNA structure  Smallest types of RNA  Highly structured  All tRNAs contain between 74 and 93 nucleotides in a single chain  Structural features: hairpin turns, regions of double helix and loops (non-hydrogen bonded portions)  Carriers of specific amino acids used for protein synthesis  Reads the codon message on mRNA and incorporates amino acid into the protein being synthesized  20 amino acid – 20 tRNA

43. tRNA – 3 o structure  To produce tertiary structure, tRNA folds into an L- shaped conformation The 3D structure of yeast tRNA for phenylalanine

44. rRNA  Ribosomal RNA, rRNA: a ribonucleic acid found in ribosomes, the site of protein synthesis –only a few types of rRNA exist in cells –ribosomes ( protein-synthesizing organelles) consist of 60 to 65% rRNA and 35 to 40% protein –in both prokaryotes and eukaryotes, ribosomes consist of two subunits, one larger than the other –analyzed by analytical ultracentrifugation –particles characterized by sedimentation coefficients, expressed in Svedberg units (S) –Sequencing of 16S RNA (small subunit of bacteria rRNA) - identification of bacteria

45. rRNA structure  Secondary & tertiary structures of rRNA display same elements as tRNAs Secondary structure for E. coli 16S rRNA.

46. mRNA  Messenger RNA, mRNA: a ribonucleic acid that carries coded genetic information from DNA to ribosomes for the synthesis of proteins – present in cells in relatively small amounts and very short-lived (less abundant form of RNA) – single stranded – biosynthesis is directed by information encoded on DNA – Synthesize from DNA, the nucleotide sequence in mRNA is similar with the 5’-3’ strand of DNA, with the exception of U replacing T 5’-3’ DNA sequence is the same with RNA sequence (complementary to 3’-5’DNA) sequence Structure: Linear polynucleotide strand

47. mRNA structure  Serves as a template for protein synthesis (Carries the transient message for protein synthesis from nuclear DNA to the ribosomes)  Move the information contained in DNA to the translation machinery  Each molecule carries the instruction for each gene (codes for one type of polypeptide product) 5’ – G G C A U U G C G C - 3’

48.  Initiation codon - codes for the 1 st amino acid in all polypeptide sequences - N-formyl methionine in prokaryotes and methionine in eukaryotes  Termination codon - do not code for an amino acid & thus signal the end of protein synthesis - Also called stop codon or nonsense codon