Analysis of organophosphate complexes provides valuable enzyme insights

A recent study led by Dr. Sussman and his team, published in Structural Biology, has shed light on the factors influencing the visualization of ligand-protein complexes using a bacterial phosphotriesterase. The study's examination of 12 crystal structures emphasized the role of protein expression tags and space group in ligand binding, underlining the need for meticulous experimental design to ensure accurate structural studies.

The research team used a bacterial phosphotriesterase as a model system to delve into the elements affecting the visualization of molecular complexes between ligands and target enzymes. They scrutinized various factors such as molecular constructs, ligands used during protein expression and purification, crystallization conditions, and space group. Organophosphates, fragments of nerve agents and insecticides, were employed as ligands. A total of 12 crystal structures of different phosphotriesterase constructs were analyzed, achieving resolutions up to 1.38 Å. Both apo and holo forms complexed with organophosphate ligands were studied. Crystals obtained from diverse conditions and space groups were examined, including those with and without N-terminal tags.

The findings revealed that protein expression tags could hinder ligand binding within the active site. Additionally, it was found that the space group of protein crystallization significantly influenced ligand visualization. The study also pointed out that crystallization precipitants could compete with or obstruct ligand binding, potentially leading to incorrect drug candidate identification. Interestingly, one co-crystallization condition facilitated defining spaces accommodating substituents on organophosphate substrates. Structures of phosphotriesterase-organophosphate complexes showed similar interaction distances between P—O bond oxygen atoms and β-Zn2+ and α-Zn2+ ions, suggesting their roles in stabilizing the substrate hydrolysis transition state.

In conclusion, Sussman's study provides valuable insights into the challenges and considerations involved in studying ligand-protein complex crystal structures. It emphasizes the need for careful experimental design and rigorous data analysis to ensure accuracy and reliability in phosphotriesterase-organophosphate structures.

The study is titled "The impact of molecular variants, crystallization conditions and the space group on ligand–protein complexes: a case study on bacterial phosphotriesterase," authored by Orly Dym et al., and published in Acta Cryst. (2023). DOI: doi.org/10.1107/S2059798323007672