Quantitative determination of low concentrations of peptides in aqueous solutions by¹⁴C-labelling

Abstract

Calibration of a counting rate method with standard solutions of¹⁴C-labelled peptides allows the accurate determination of 1·10⁻⁶ to 1·10⁻²M concentrations of peptides in aqueous solution. By the use of this method and the equilibrium dialysis technique it is shown that binding constants of different peptides to DNA that differ by as little as ΔK=|0.3|mM⁻¹ can be measured. Statistical error considerations show that such a result is significant. The use of colored complexes formed between 2,4-dinitrofluorobenzene and LYS-LYS and LYS-TRYP-LYS enabled the affinity constants of these peptides with calf-thymus DNA to be determined. The advantages of the determination of peptide concentrations, by the use of radioactive counting, over colorimetric methods as well as the importance of this highly sensitive method for the measurement of peptide concentrations and hence binding constants are discussed.