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Protein interactions in solution characterized by light and neutron scattering: comparison of lysozyme and chymotrypsinogen.

ABSTRACT: The effects of pH and electrolyte concentration on protein-protein interactions in lysozyme and chymotrypsinogen solutions were investigated by static light scattering (SLS) and small-angle neutron scattering (SANS). Very good agreement between the values of the virial coefficients measured by SLS and SANS was obtained without use of adjustable parameters. At low electrolyte concentration, the virial coefficients depend strongly on pH and change from positive to negative as the pH increases. All coefficients at high salt concentration are slightly negative and depend weakly on pH. For lysozyme, the coefficients always decrease with increasing electrolyte concentration. However, for chymotrypsinogen there is a cross-over point around pH 5.2, above which the virial coefficients decrease with increasing ionic strength, indicating the presence of attractive electrostatic interactions. The data are in agreement with Derjaguin-Landau-Verwey-Overbeek (DLVO)-type modeling, accounting for the repulsive and attractive electrostatic, van der Waals, and excluded volume interactions of equivalent colloid spheres. This model, however, is unable to resolve the complex short-ranged orientational interactions. The results of protein precipitation and crystallization experiments are in qualitative correlation with the patterns of the virial coefficients and demonstrate that interaction mapping could help outline new crystallization regions.

SUBMITTER: Velev OD

PROVIDER: S-EPMC1299943 | biostudies-other | 1998 Dec

REPOSITORIES: biostudies-other

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Protein interactions in solution characterized by light and neutron scattering: comparison of lysozyme and chymotrypsinogen.

Velev O D OD Kaler E W EW Lenhoff A M AM

Biophysical journal 19981201 6

The effects of pH and electrolyte concentration on protein-protein interactions in lysozyme and chymotrypsinogen solutions were investigated by static light scattering (SLS) and small-angle neutron scattering (SANS). Very good agreement between the values of the virial coefficients measured by SLS and SANS was obtained without use of adjustable parameters. At low electrolyte concentration, the virial coefficients depend strongly on pH and change from positive to negative as the pH increases. All ...[more]

PMID: 9826592

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Project description:Protein-protein interactions were investigated for ?-chymotrypsinogen by static and dynamic light scattering (SLS and DLS, respectively), as well as small-angle neutron scattering (SANS), as a function of protein and salt concentration at acidic conditions. Net protein-protein interactions were probed via the Kirkwood-Buff integral G22 and the static structure factor S(q) from SLS and SANS data. G22 was obtained by regressing the Rayleigh ratio versus protein concentration with a local Taylor series approach, which does not require one to assume the underlying form or nature of intermolecular interactions. In addition, G22 and S(q) were further analyzed by traditional methods involving fits to effective interaction potentials. Although the fitted model parameters were not always physically realistic, the numerical values for G22 and S(q ? 0) were in good agreement from SLS and SANS as a function of protein concentration. In the dilute regime, fitted G22 values agreed with those obtained via the osmotic second virial coefficient B22 and showed that electrostatic interactions are the dominant contribution for colloidal interactions in ?-chymotrypsinogen solutions. However, as protein concentration increases, the strength of protein-protein interactions decreases, with a more pronounced decrease at low salt concentrations. The results are consistent with an effective "crowding" or excluded volume contribution to G22 due to the long-ranged electrostatic repulsions that are prominent even at the moderate range of protein concentrations used here (<40 g/L). These apparent crowding effects were confirmed and quantified by assessing the hydrodynamic factor H(q ? 0), which is obtained by combining measurements of the collective diffusion coefficient from DLS data with measurements of S(q ? 0). H(q ? 0) was significantly less than that for a corresponding hard-sphere system and showed that hydrodynamic nonidealities can lead to qualitatively incorrect conclusions regarding B22, G22, and static protein-protein interactions if one uses only DLS to assess protein interactions.

Project description:The binding interactions of three gemini surfactants having different spacer groups (12-4-12, 12-8-12, and 12-4(OH)-12) with a high concentration (150 μM) of bovine serum albumin (BSA) at various regions of binding isotherms have been studied by means of steady-state fluorescence and fluorescence anisotropy, time-correlated single-photon counting fluorescence of trans-2-[4-(dimethylamino)styryl]benzothiazole, small-angle neutron scattering (SANS), and dynamic light scattering (DLS) measurements. The fluorescence resonance energy transfer phenomenon between the twisted intramolecular charge transfer fluorescent molecule, trans-2-[4-(dimethylamino)styryl]benzothiazole as an acceptor, and tryptophan 213 (Trp-213) of BSA as a donor has been successfully used to probe the binding interactions of gemini surfactants with protein at all regions of binding isotherms. The increasing order of energy transfer efficiency at a higher concentration range of surfactants is 12-8-12 > 12-4-12 > 12-4(OH)-12. Stronger binding of micelles of gemini surfactant molecules having a comparatively more hydrophobic spacer group with the hydrophobic segments of the protein results in closer approach of trans-2-[4-(dimethylamino)styryl]benzothiazole molecules solubilized in micelles to Trp-213. The average excited-state lifetimes become shorter with a trend of increase in contribution from the fast component and decrease in contribution from the slow component to the decay with increasing concentration of a surfactant. The nonradiative rate constant of trans-2-[4-(dimethylamino)styryl]benzothiazole increases with increasing concentration of a surfactant because the average microenvironment around it in protein-surfactant aggregates is more polar as compared to that in native protein. SANS and DLS measurements were carried out for the study of the structural deformations in the protein, on enhancement of the concentration of the gemini surfactants. The necklace and bead model has been used for the analysis of SANS data for the protein-surfactant complexes. At a higher concentration range, 12-8-12 and 12-4-12 have a slightly smaller fractal dimension and a larger correlation length as compared to 12-4(OH)-12. DLS data show that the increasing order of hydrodynamic diameter for the complexes of protein with three gemini surfactants in their high concentration range is 12-4(OH)-12 < 12-4-12 < 12-8-12.

Dataset Information

Protein interactions in solution characterized by light and neutron scattering: comparison of lysozyme and chymotrypsinogen.

Publications

Protein interactions in solution characterized by light and neutron scattering: comparison of lysozyme and chymotrypsinogen.

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