Native State of Complement Protein C3d Analysed via Hydrogen Exchange and Conformational Sampling

Area of Science:

• Biochemistry
• Structural Biology
• Computational Biology

Background:

• Hydrogen/deuterium exchange detected by mass spectrometry (HDX-MS) is crucial for studying protein structure and dynamics.
• Interpreting HDX-MS data traditionally relies on crystal structures, but conformational ensembles from molecular dynamics simulations are proposed as more accurate.
• The complement protein C3d serves as a model system for evaluating HDX-MS data interpretation methods.

Purpose of the Study:

• To analyze complement protein C3d using HDX-MS.
• To evaluate different methodologies for interpreting HDX-MS data by computationally replicating experimental results.
• To identify the most suitable structural representation for accurate HDX-MS data interpretation.

Main Methods:

• Performed an HDX-MS experiment on complement protein C3d.
• Utilized an existing prediction model to derive HDX-MS data from protein structures.
• Evaluated crystal structures, molecular dynamics simulations, and coarse-grained conformational sampling for data interpretation.
• Sought a C3d conformation or ensemble that computationally replicates experimental HDX-MS data.

Main Results:

• Crystal structures were confirmed as inadequate for interpreting HDX-MS data.
• Molecular dynamics-derived conformational ensembles did not consistently provide satisfactory interpretations.
• Coarse-grained conformational sampling of C3d yielded a conformation capable of replicating and refining the experimental HDX-MS data.

Conclusions:

• Crystal structures are not optimal for HDX-MS data interpretation.
• Molecular dynamics simulations may not always yield satisfactory conformational ensembles for HDX-MS analysis.
• Coarse-grained conformational sampling offers a promising approach for accurate HDX-MS data interpretation and refinement, as demonstrated with complement protein C3d.