pH-Dependent PdS₃ Site in α₃DIV Revealed by Single-Molecule Force Spectroscopy

Area of Science:

• Biochemistry
• Toxicology
• Structural Biology

Background:

• Lead is a toxic metal that exerts harmful effects by interacting with proteins.
• Lead's thiophilic nature drives its high affinity for thiol-rich proteins, forming stable lead-sulfur (Pb-S) bonds.
• Understanding Pb-S bonds is crucial for elucidating lead toxicity mechanisms.

Purpose of the Study:

• To investigate the formation and characteristics of Pb-S bonds in a model protein.
• To characterize the Pb-S bonds within the de novo designed protein α3DIV, which binds Pb(II) via three cysteine residues.

Main Methods:

• Utilized atomic force microscopy-based single-molecule force spectroscopy (AFM-SMFS).
• Employed a de novo designed protein (α3DIV) engineered to bind Pb(II) through its cysteine residues.

Main Results:

• Confirmed the formation of a triangular pyramidal PbS3 site in α3DIV.
• Detected specific Pb-S bond rupture signals, including the previously unobserved Pb-S(Cys67) bond.
• Quantified the pH-dependent weakening of Pb-S bond strength, observing dissociation of the PbS3 site at pH 4.5.

Conclusions:

• AFM-SMFS is effective in characterizing Pb-S bonds in proteins at the single-molecule level.
• The study provides new insights into the structural and dynamic properties of lead-protein interactions.
• Understanding the pH-dependent stability of Pb-S bonds is important for predicting lead's behavior in biological systems.