RESUMO
In macromolecular crystallization, success is often dependent on the pH of the experiment. However, little is known about the pH of reagents used, and it is generally assumed that the pH of the experiment will closely match that of any buffering chemical in the solution. We use a large dataset of experimentally measured solution pH values to show that this assumption can be very wrong and generate a model that can be used to successfully predict the overall solution pH of a crystallization experiment. Furthermore, we investigate the time dependence of the pH of some polyethylene glycol polymers widely used in protein crystallization under different storage conditions.
RESUMO
The Protein Data Bank (PDB) is the largest available repository of solved protein structures and contains a wealth of information on successful crystallization. Many centres have used their own experimental data to draw conclusions about proteins and the conditions in which they crystallize. Here, data from the PDB were used to reanalyse some of these results. The most successful crystallization reagents were identified, the link between solution pH and the isoelectric point of the protein was investigated and the possibility of predicting whether a protein will crystallize was explored.
Assuntos
Bases de Dados de Proteínas , Estatística como Assunto , Sulfato de Amônio/química , Cristalização , Concentração de Íons de Hidrogênio , Ponto Isoelétrico , Polietilenoglicóis/químicaRESUMO
MOTIVATION: The identification of suitable conditions for crystallization is a rate-limiting step in protein structure determination. The pH of an experiment is an important parameter and has the potential to be used in data-mining studies to help reduce the number of crystallization trials required. However, the pH is usually recorded as that of the buffer solution, which can be highly inaccurate. RESULTS: Here, we show that a better estimate of the true pH can be predicted by considering not only the buffer pH but also any other chemicals in the crystallization solution. We use these more accurate pH values to investigate the disputed relationship between the pI of a protein and the pH at which it crystallizes. AVAILABILITY AND IMPLEMENTATION: Data used to generate models are available as Supplementary Material. CONTACT: julie.wilson@york.ac.uk SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Assuntos
Cristalização , Proteínas/química , Concentração de Íons de Hidrogênio , Ponto IsoelétricoRESUMO
The crystallization of proteins is dependent on the careful control of numerous parameters, one of these being pH. The pH of crystallization is generally reported as that of the buffer; however, the true pH has been found to be as many as four pH units away. Measurement of pH with a meter is time-consuming and requires the reformatting of the crystallization solution. To overcome this, a high-throughput method for pH determination of buffered solutions has been developed with results comparable to those of a pH meter.