Characterization of the acid/base and redox chemistry of phytochelatin analogue peptides.

The phytochelatins are a family of polydisperse, thiol-rich peptides that are synthesized by plants in response to exposure to heavy metals. The amino acid sequence of the phytochelatin peptides is (gamma-glutamyl-cysteinyl)n-glycine, where n typically ranges from 2 to 5. In the first phase of a program to characterize the coordination chemistry of the phytochelatins with heavy metals, the phytochelatin analogue peptides acetyl(gamma-glutamyl-cysteinyl)n-glycine amide (Ac-(gamma-Glu-Cys)n-NH2, n = 2-6) have been synthesized by solid-phase peptide synthesis methods and characterized by 1H NMR spectroscopy. The 1H NMR spectra of the analogue peptides were completely assigned by using band-selective homonuclear-decoupled (BASHD) two-dimensional NMR experiments to achieve spectral resolution. The acid dissociation constant of each cysteine residue in each peptide was determined from chemical shift-pH titration data for the CalphaH protons of the cysteine residues. The resonances for the CalphaH protons were resolved in BASHD-total correlation spectroscopy spectra that were measured as a function of pH. The pKA values for a given thiol group depend on the position of the cysteine residue in the sequence, with the thiol group of the cysteine residue attached to the C-terminal glycine being the most acidic. The pKA values also depend on the size of the peptide, increasing as the size, and thus the negative charge, of the peptide increases. The redox potential for oxidation of the two thiol groups of Ac(gamma-Glu-Cys)2-NH2 to form an intramolecular disulfide bond was also determined by measuring the equilibrium constant for its thiol/disulfide exchange reaction with glutathione.