Cd2+ and Pb2+ complexation by glutathione and the phytochelatins

Abstract

Phytochelatins or PCn, (γGlu-Cys)_n-Gly, and their glutathione (GSH) precursor are thiol-rich peptides that play an important role in heavy metal detoxification in plants and microorganisms. Complex formation between Cd²⁺ and Pb²⁺ and GSH or PCn (n = 2, 4 and 6) are investigated by microcalorimetry, absorption spectrophotometry and T-jump kinetics. Complex formation with Pb²⁺ or Cd²⁺ is exothermic, and induces ligand metal charge transfer bands in UV absorption spectral range, which implies the formation of a coordination bond between the metal and the thiol groups of the phytochelatins. Absorption spectra and microcalorimetry experiments allow the determination of the affinity constants and the stoichiometry of the complexes. We show that the three PCn interact with Pb²⁺ to form the 1:1 and 2:1 M:L complexes, with similar affinity constants (log K₁₁^Pb∼4.6, log K₂₁^Pb∼11.4). These affinities are independent of the number of thiols and are, moreover, lower than those determined for complex formation with Cd²⁺. On the other hand, with Cd²⁺, PC2-Cd, PC2-Cd₂, (PC2)₃-Cd₂, PC4-Cd, PC4-Cd₂, PC6-Cd, (PC6)₂-Cd₃ and PC6-Cd₃ complexes are detected. Furthermore, for PC4-Cd, the 1:1 complex is the most stable: affinity constant (log K₁₁^Cd∼7.5). Kinetic studies indicate that complex formation between Cd²⁺ and GSH occurs in the ms range; direct rate constant k_obs = (6.8 ± 0.3) 10⁶ M^-1 s^-1 and reverse rate constant k_-obs = 340 ± 210 s^-1. Thus, when encapsulated in a silica matrix, PCn can be good candidates for heavy metal detection.

Keywords: Fast kinetics; Metal interactions; Microcalorimetry; Phytoremediation; Spectroscopy.