BIOCH200 Lecture Notes - Lecture 6: Nitrogenous Base, Nucleic Acid Thermodynamics, Deamination
Nucleic Acids
❏Polymers of nucleotides linked by 3’-5’ phosphodiester bonds.
❏Nucleotides are written from 5’ to 3’ by convention.
❏5’ end has a free phosphate group → not involved in a phosphodiester bond
❏3’ end has a free OH group not involved in phosphodiester bond.
❏Net Charge → Negative due to phosphates
❏RNA is more polar because of OH group (3 more H-bonds)
❏DNA backbone → Sugar and Phosphate → Polar
Representation of Primary Structures of Nucleic Acids
1. 5’-ATG-3’ → DNA due to presence of T.
2. 5’-ACG-3’ → likely RNA because DNA would be dAdCdG
3. 5-UCG-3’ → RNA due to presence of U
4. ACGC → By convention A is 5’ and C is 3’ → RNA because DNA would be dAdCdGdC
5. ACGCT → pApCpGpCpT → phosphodiester linkage, 5’ is A, 3’ is T, DNA due to presence of T.
Polynucleotides
❏Mononucleotide → one nitrogen base, sugar and phosphate
❏Dinucleotide → 2 bases, 2 sugars and 2 phosphates.
❏Oligonucleotide → 20-50
❏Polynucleotides → Large number of nucleic acids.
Hydrolysis of Phosphodiester bonds
❏Could occur on either side
❏You may also undergo alkaline hydrolysis of phosphodiester bonds where pH is typically greater
than 9 and the compound has an OH group.
❏RNa is more susceptible to alkaline hydrolysis due to OH group. DNA is more stable due
to lack of OH group.
Cytosine Deamination
❏Less serious in RNA → short life span
❏More serious in DNA → Larger life span.
Properties of the Bases (purines/pyrimidines)
❏Heterocyclic and Aromatic (electron delocalization)
❏Allow for Abs UV light → Max. ~260nm.
❏Planar, slight pucker in purine bases
❏Poorly soluble, and largely hydrophobic with some polar groups
❏Able to form H-bonds.
❏Sugar phosphate backbone is polar.
Electron delocalization allows bases to absorb UV light
❏Ratio of A260/A280 tells us the purity of DNA. At approx 1.8-1.9 it is Pure DNA.
❏If the Ratio is lower than 1.8-1.9 there is probably protein in the sample because it has a max
absorbance at ~280nm.
❏Determine amount DNA by graphing [DNA] in relation to A260nm. As [DNA] increases A260
should increase.
❏ssDNA absorbs higher than dsDNA
❏DNA and RNA are very similar that you can’t determine the two based on ratio.
DNA Structure
❏Erwin Chargaff → A=T, C=G in dsDNA
Document Summary
Polymers of nucleotides linked by 3"-5" phosphodiester bonds. Nucleotides are written from 5" to 3" by convention. 5" end has a free phosphate group not involved in a phosphodiester bond. 3" end has a free oh group not involved in phosphodiester bond. Net charge negative due to phosphates. Rna is more polar because of oh group (3 more h-bonds) Dna backbone sugar and phosphate polar. Mononucleotide one nitrogen base, sugar and phosphate. Dinucleotide 2 bases, 2 sugars and 2 phosphates. Polynucleotides large number of nucleic acids. You may also undergo alkaline hydrolysis of phosphodiester bonds where ph is typically greater than 9 and the compound has an oh group. Rna is more susceptible to alkaline hydrolysis due to oh group. Dna is more stable due to lack of oh group. Less serious in rna short life span. More serious in dna larger life span. Allow for abs uv light max. Poorly soluble, and largely hydrophobic with some polar groups.