Dataset Information

Determination of the phase diagram for soluble and membrane proteins.

ABSTRACT: Methods to efficiently determine the phase behavior of novel proteins have the potential to significantly benefit structural biology efforts. Here, we present protocols to determine both the solubility boundary and the supersolubility boundary for protein/precipitant systems using an evaporation-based crystallization platform. This strategy takes advantage of the well-defined rates of evaporation that occur in this platform to determine the state of the droplet at any point in time without relying on an equilibrium-based end point. The dynamic nature of this method efficiently traverses phase space along a known path, such that a solubility diagram can be mapped out for both soluble and membrane proteins while using a smaller amount of protein than what is typically used in optimization screens. Furthermore, a variation on this method can be used to decouple crystal nucleation and growth events, so fewer and larger crystals can be obtained within a given droplet. The latter protocol can be used to rescue a crystallization trial where showers of tiny crystals were observed. We validated both of the protocols to determine the phase behavior and the protocol to optimize crystal quality using the soluble proteins lysozyme and ribonuclease A as well as the membrane protein bacteriorhodopsin.

SUBMITTER: Talreja S

PROVIDER: S-EPMC2848416 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Determination of the phase diagram for soluble and membrane proteins.

Talreja Sameer S Perry Sarah L SL Guha Sudipto S Bhamidi Venkateswarlu V Zukoski Charles F CF Kenis Paul J A PJ

The journal of physical chemistry. B 20100401 13

Methods to efficiently determine the phase behavior of novel proteins have the potential to significantly benefit structural biology efforts. Here, we present protocols to determine both the solubility boundary and the supersolubility boundary for protein/precipitant systems using an evaporation-based crystallization platform. This strategy takes advantage of the well-defined rates of evaporation that occur in this platform to determine the state of the droplet at any point in time without relyi ...[more]

PMID: 20235520

Similar Datasets

Project description:This article presents the results of a detailed analysis of helix-helix interactions in membrane and soluble proteins. A data set of interacting pairs of helices in membrane proteins of known structure was constructed and a structure alignment algorithm was used to identify pairs of helices in soluble proteins that superimpose well with pairs of helices in the membrane-protein data set. Most helix pairs in membrane proteins are found to have a significant number of structural homologs in soluble proteins, although in some cases, primarily involving irregular helices, no close homologs exist. An analysis of geometric relationships between interacting helices in the two sets of proteins identifies some differences in the distributions of helix length, interfacial area, packing angle, and distance between the polypeptide backbones. However, a subset of soluble-protein helix pairs that are close structural homologs to membrane-protein helix pairs exhibits distributions that mirror those observed in membrane proteins. The larger average interface size and smaller distance of closest approach seen for helices in membrane proteins appears due in part to a relative enrichment of alanines and glycines, particularly as components of the AxxxA and GxxxG motifs. It is argued that membrane helices are not on average more tightly packed than helices in soluble proteins; they are simply able to approach each other more closely. This enables them to interact over longer distances, which may in turn facilitate their remaining in contact over much of the width of the lipid bilayer. The close structural similarity seen between some pairs of helices in membrane and soluble proteins suggests that packing patterns observed in soluble proteins may be useful in the modeling of membrane proteins. Moreover, there do not appear to be fundamental differences between the magnitude of the forces that drive helix packing in membrane and soluble proteins, suggesting that strategies to make membrane proteins more soluble by mutating surface residues are likely to encounter success, at least in some cases.

Dataset Information

Determination of the phase diagram for soluble and membrane proteins.

Publications

Determination of the phase diagram for soluble and membrane proteins.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets