Lecture6 - Lecture 6 1/16/09 Assignment: Segel, Pages...

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Lecture 6 1/16/09 Outline Isoelectric point Ion-exchange chromatography Amino acid sequencing Amino-terminal amino acid determination Carboxyl-terminal amino acid determination Specific fragmentation of peptides Assignment: Segel, Pages 141-142: Problems 1, 2, 3 and 5 Background Reading: Segel, Page 94
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H H 2 N H 2 N NH pK a1 = 2.2 pK a2 = 9.0 pK a3 = 12.5 +2 +1 0 -1 Isoelectric point Net charge: The isoelectric point of an amino acid or peptide is the pH at which it carries no net charge. It is often abbreviated as p I p I of arginine = pK a2 + pK a3 2 = 10.75
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H H H H At pH 3, the amino acids shown below would be primarily in their fully protonated forms. p I : 2.98 5.97 9.74 Order of elution from cation-exchange column: Aspartate, Glycine, Lysine
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Ion-exchange chromatography Cation-exchange using a resin with sulfonic acid groups attached. SO 3 - SO 3 - SO 3 - CH 2 CH CH CH 2 n Dowex-50 The positive charges of the amino acids will bind to the resin by electrostatic forces.
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Lecture6 - Lecture 6 1/16/09 Assignment: Segel, Pages...

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