Protein crystallization by design: chymotrypsinogen without precipitants

Area of Science:

• Biochemistry
• Structural Biology
• Crystallography

Background:

• Protein crystallization typically relies on empirical screening.
• Existing bovine chymotrypsinogen A structures were determined using ethanol-containing solutions.

Purpose of the Study:

• To develop and apply a systematic predictive procedure for protein crystallization.
• To obtain high-resolution X-ray structure of bovine chymotrypsinogen A using novel conditions.

Main Methods:

• Measurements of osmotic second virial coefficient to determine suitable solvent conditions.
• Ultracentrifugal crystallization for approximately 30 hours without precipitants.
• X-ray structure determination to 3 Å resolution.

Main Results:

• Data-quality crystals of bovine chymotrypsinogen A were produced using buffered NaCl solutions.
• A refined X-ray structure was obtained with high map quality, defining most protein atoms.
• Crystallization occurred in the tetragonal space group P4(1)2(1)2.

Conclusions:

• A systematic, predictive approach to protein crystallization is feasible.
• This method expands the range of successful crystallization conditions and can yield new crystal forms.
• Observed minor structural differences suggest potential effects of prior crystallization conditions.