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Thallium counterion distribution in cubic insulin crystals determined from anomalous x-ray diffraction data
1Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, MA 02254-9110.
Summary
This study used anomalous scattering diffraction to map thallium ion (Tl+) distribution around insulin crystals. Six Tl+ binding sites were identified, revealing cation interactions with protein and water molecules.
Area of Science:
- Biophysics
- Structural Biology
- Crystallography
Background:
- Understanding cation distribution is crucial for protein structure and function.
- Insulin's charge varies with pH, affecting counterion interactions.
Purpose of the Study:
- To determine the precise locations and characteristics of monovalent cations around insulin crystals.
- To investigate how pH influences cation binding to insulin.
Main Methods:
- Anomalous scattering diffraction data collection from thallium (Tl)-containing cubic insulin crystals.
- Calculation of anomalous scattering difference maps using Bijvoet reflection pairs.
- Analysis of Tl+ ion distribution at pH 8 and pH 10.5.
Main Results:
- Six thallium (Tl+) sites were identified, including two previously known and four newly characterized mobile sites.
- Three Tl+ sites interact with protein carbonyl dipoles, not negatively charged groups.
- Tl+ ions interact with both protein atoms and water molecules, with occupancies influenced by ion interactions and protein flexibility.
Conclusions:
- Cation distribution is complex, influenced by protein structure and pH-dependent net charge.
- At pH 10.5, a higher protein net charge necessitates more disordered counterions than observed.
- Anomalous scattering difference maps effectively visualize specific cation binding sites.