Copper trafficking: the solution structure of Bacillus subtilis CopZ

Area of Science:

• Biochemistry
• Structural Biology
• Microbiology

Background:

• A copper-transport CopZ protein homolog was identified in Bacillus subtilis, sharing homology and a metal-binding motif with Enterococcus hirae CopZ.
• The Bacillus subtilis CopZ protein (73 amino acids) was expressed and characterized.

Purpose of the Study:

• To determine the solution structure of the copper(I)-bound Bacillus subtilis CopZ protein.
• To compare the structural properties of Bacillus subtilis CopZ with Enterococcus hirae CopZ.
• To investigate the role and conserved residues in copper-binding proteins.

Main Methods:

• Nuclear Magnetic Resonance (NMR) spectroscopy was used to determine the solution structure of the copper-bound protein.
• NMR data were analyzed using nuclear Overhauser effects and dihedral angles for structural calculations.
• The structure of the copper-free apoprotein was also investigated using NMR.

Main Results:

• The Bacillus subtilis CopZ protein binds copper(I) under reducing conditions and is fully folded in both copper-bound and copper-free forms.
• The solution structure of the copper-bound form was determined, revealing a well-defined structure.
• NMR data indicated that the apoprotein also maintains a folded state with intact secondary structure, though some loop disorder was observed.

Conclusions:

• Bacillus subtilis CopZ is a folded copper-binding protein, distinct in its behavior from Enterococcus hirae CopZ, possibly due to stabilizing agents like dithiothreitol.
• Conserved residues and the metal-binding loop are crucial for copper binding.
• The study provides insights into the function and structural characteristics of CopZ proteins.