Protein production and crystallization at SECSG -- an overview

Area of Science:

• Structural biology
• Genomics
• Biochemistry

Background:

• High-throughput pipelines are crucial for structural genomics.
• Protein Data Bank (PDB) requires new protein structures.
• The Southeast Collaboratory for Structural Genomics (SECSG) addresses this need.

Purpose of the Study:

• To describe the SECSG's automated, high-throughput pipelines for protein production and crystallization.
• To detail a two-tiered approach for efficient protein structure determination.
• To present strategies for target rescue in structural biology.

Main Methods:

• Implemented automated, high-throughput pipelines for protein production and crystallization.
• Utilized a two-tiered approach: Tier-1 for initial production, Tier-2 for target rescue.
• Screened over 40 proteins per week using the crystallization pipeline.

Main Results:

• Developed efficient pipelines for producing and crystallizing proteins from Pyrococcus furiosus, Caenorhabditis elegans, and human targets.
• Tier-1 focused on novel Pfam families lacking PDB structures.
• Tier-2 successfully rescued proteins that failed to crystallize or yielded poor-quality crystals in Tier-1.

Conclusions:

• The described two-tiered, automated approach significantly enhances efficiency in protein production and crystallization.
• SECSG's pipelines accelerate the structure determination process for underrepresented protein families.
• This methodology provides a robust framework for advancing structural genomics initiatives.