Implementation of semi-automated cloning and prokaryotic expression screening: the impact of SPINE

The technology output from the Structural Proteomics In Europe (SPINE) consortium for cloning and expression in E. coli is reviewed.

Eukaryotic expression: developments for structural proteomics

Strategies and experience resulting from the use of eukaryotic expression systems in SPINE are described.

The impact of protein characterization in structural proteomics

Characteristics such as the purity of target proteins are key factors that can influence the success of structure determination. Here, the application and efficacy of various quality-assessment methodologies are discussed.

SPINE high-throughput crystallization, crystal imaging and recognition techniques: current state, performance analysis, new technologies and future aspects

The introduction of automation and minimization into the crystallization pipeline has improved the yield of high-quality crystals for structural determination using more reproducible experimental protocols.

NMR in the SPINE Structural Proteomics project

New hardware and software developments for protein structure determination by NMR are described. These techniques allow faster, more automated solution-structure determination and more efficient data utilization.

High-throughput sample handling and data collection at synchrotrons: embedding the ESRF into the high-throughput gene-to-structure pipeline

Automated data collection for macromolecular crystallography at beamlines with protocols for automatic instrumentation improves the efficient use of available beamtime. These developments were a result of a collaboration between the main partners of SPINE and members of other similar European and national initiatives.

SPINE workshop on automated X-ray analysis: a progress report

The results of a workshop on the automation of protein crystal structure solution organized by the Structural Proteomics In Europe (SPINE) consortium are reported.

SPINE bioinformatics and data-management aspects of high-throughput structural biology

This article reviews the infrastructure of software that has been developed under the SPINE initiative for high-throughput protein-structure analysis covering target selection and prototype LIMS for protein production through to structure annotation, with contributions to the management of a large-scale crystallization set-up and structure determination.

Application of the use of high-throughput technologies to the determination of protein structures of bacterial and viral pathogens

Progress by the SPINE consortium on the determination of structures of proteins of bacterial and viral pathogens is outlined, with particular reference to the impact of high-throughput technologies.

First steps towards effective methods in exploiting high-throughput technologies for the determination of human protein structures of high biomedical value

Progress in the determination of structures of human proteins by the Structural Proteomics In Europe (SPINE) consortium is outlined with highlights indicating the impact of high-throughput technologies.

Recombinant protein expression and solubility screening in Escherichia coli: a comparative study

This article presents a comparative analysis of the standard methods for small-scale screening of recombinant protein expression/solubility implemented in laboratories of the Structural Proteomics In Europe (SPINE) consortium.

GRETA, a new multifermenter system for structural genomics and process optimization

A multifermentation system, GRETA, has been developed which specifically accommodates features that respond to the needs of future structural genomics programmes. The GRETA multifermenter provides a robust platform for streamlined parallel fermentation, requiring minimal manual intervention, while at the same time providing a versatile fermentation platform for process optimization.

Co-expression of protein complexes in prokaryotic and eukaryotic hosts: experimental procedures, database tracking and case studies

The design of a new system for multi-expression in E. coli and a database for co-expression data are described. Lessons are drawn from case studies using these and other strategies for co-expression and analysis of protein complexes.

A time- and cost-efficient system for high-level protein production in mammalian cells

A simple, fast and affordable method for transient protein expression in mammalian cells is presented. It combines several features of interest for X-ray crystallography such as high protein yield, straightforward purification, selenomethinonine incorporation and control of N-linked glycosylation.

Automation of sample mounting for macromolecular crystallography

A sample holder standard for use with robotic sample changers is defined. The standard includes a system for sample identification, tracking and management of data flow in a macromolecular structure-determination pipeline. A robotic sample changer designed for use with the sample standard is described.

Honing the in silico toolkit for detecting protein disorder

The current state of methods for the in silico prediction of natively disordered regions of proteins is discussed in the context of the potential for improvement and application of these predictions in structural proteomics.

Application of high-throughput technologies to a structural proteomics-type analysis of Bacillus anthracis

The current state of high-throughput technologies for protein production and structure determination is assessed in relation to progress on the SPINE structural proteomics project on proteins from B. anthracis.

Structural genomics of the Epstein–Barr virus

For a structural genomics project on Epstein–Barr virus, 23 proteins were cloned into E. coli for protein expression. The structures of four have been solved by X-ray crystallography.