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Unformatted text preview: 542 2.772/20.110 Problem Set #7 Solutions 1.) P19.8) An important biological application of absorption spectroscopy is the determination of the concentrations of solutions of nucleic acids. The * electronic transitions within the purine and pyrimidine bases of nucleic acids have absorption maxima near a wavelength of 260 nm. Assume the extinction coefficient of a nucleic acid is 1.00 10 4 M 1 cm 1 at 260 nm. If the concentration of a nucleic acid solution is 5.00 10 4 M, calculate the absorbance of this solution in a 1.00 cm cell at 260 nm. l c I I log A = = 1.00cm M 10 5 cm M 1 10 1 A 4- 4 260 = 5 A 260 = 2.) P19.9) The absorbance of a nucleic acid solution at 260 nm is called A 260 . The OD (i.e., optical density) is the amount of nucleic acid in a volume of 1.00 mL in a 1.00-cm path length cell for which A 260 =1.00. How many moles of nucleotide are contained in a 1.00 mL solution of a double-stranded nucleotide for which A 260 = 2.50, assuming the extinction coefficient per nucleotide is 7.000 10 3 M 1 cm 1 . Express this quantity in ODs. Assume a 1.00 cm path length. Solving l c I I log A = = for the concentration, c, gives: l A c = Therefore the concentration of nucleic acid in the solution is: Chapter 19/Electronic Spectroscopy 543 cm cm M 1 10 7 2.5 c 1 1 3 =-- l mol 4 10 5714 . 3 c- = Consequently, 1 ml of nucleic acid solution contains 3.5714 10-7 moles. This concentration corresponds to an A = 2.5, 3.) P19.10) Because of interactions between transition dipoles of the constituent nucleotides, the extinction coefficient for a single strand polynucleotide is not simply the sum of the extinction coefficients for the individual nucleotides. These dipole-dipole interactions depend on 1/ r 3 where r is the distance between bases, so for the purpose of calculating the extinction coefficient for a single-stranded polynucleotide, only nearest neighbor interactions need be considered. For a hypothetical polynucleotide strand GpCpUp ... ApG the extinction coefficient is ( GpCpUp ... ApG ) = 2 [ ( GpC ) + ( CpU ) + ( ApG )] [ ( Cp ) + ( Up ) + ... ( Ap )] Interacting nucleotides pairs in a single strand are indicated by XpY, where p represents the phosphate group that joins the nucleotides X and Y. In the equation above ( ApG ), ( ApC ), etc., are extinction coefficients for component dinucleotide phosphates per mole of nucleotide. Hence they are counted twice, which accounts for the 2 in the expression above. To correct for this fact, they are counted twice, which accounts for the 2 in the expression above....
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This note was uploaded on 11/12/2008 for the course 20 20.110 taught by Professor Griffith during the Fall '08 term at MIT.
- Fall '08