Shape Determination of Proteins in Dilute Solutions with SAXS Measurements

Topics Covered

Introduction
Measurements and Results
Conclusion
About Xenocs

Introduction

Small angle X-ray scattering (SAXS) is becoming extremely popular as a method for studying biological systems, and specifically for protein solutions. By envelope reconstruction of the studied protein, SAXS measurements enable determination of macromolecular shape conformation.

The collection of SAXS data was performed for the standard protein Lysozyme, using the Xenocs Nano-inXider. The radius of gyration (Rg) and the pair distance distribution function (PDDF) were determined from this data.

Measurements and Results

Measurements were done on 1.5, 3.0 and 5.0mg/ml concentration solutions with the Xenocs Capillary Flow Cell, using the buffer 40mM acetic acid 50mM NaCIpH 4.0.

Using the PRIMUS1 software, the structural parameter Rg results were taken. The obtained values for each concentration with a range of exposure times are given in Table 1. The values are consistent with the synchrotron data2 of Rg = 1.43nm. 10 min short exposure times are enough to determine these essential structural parameters.

Table 1. Radius of gyration of Lysozyme depending on the concentration and exposure time

c (mg/ml) Exposure time Guinier Rg (error) AutoRg1
1.5 mg/ml 10 min
30 min
1.36 nm (0.34)
1.41 nm (0.04)
3.0 mg/ml 10 min
30 min
1.43 nm (0.22)
1.40 nm (0.15)
5.0 mg/ml 10 min
30 min
1.38 nm (0.14)
1.39 nm (0.02)

The GNOM1 software was used to calculate the PPDF p(r). In Figure 1, it can be observed that the curves obtained from different concentrations overlap. Consistent data can thus be collected from low concentration measurements.

Figure 1. Pair-distance distribution function for c = 1.5, 3.0 and 5.0mg/ml. Exposure time = 30min.

Figure 2 shows the comparison between two exposure times for 5mg/ml concentration. The curves almost overlap, which is an indication of how a short exposure time of 10min is enough to provide relevant data.

Figure 2. Pair-distance distribution function for c = 5mg/ml. Exposure times = 10 and 30min.

Conclusion

Xenocs' clean-beam technology3 is completely integrated in the Nano-inXider, enabling precise bio-macromolecular studies of highly diluted systems. In addition, the use of the Xenocs Low Noise Flow Cell brings down the container scattering to push further the limits of BioSAXS measurements in the lab.

References

  1. Petoukhov et al., J. Appl. Cryst., 2007,40, s223-s2282
  2. Svergun et al, J. Appl. Cryst., 2005, 38, 555–558
  3. Xeuss Technical Note – Demonstration of High Signal-to-Noise Ratio

About Xenocs

Since its creation in 2000, Xenocs has been dedicated to offering its customers innovative solutions for X-ray characterization of nanomaterials and nanostructures.

Xenocs has over 14 years' experience in the development and production of high performance instruments for characterizing the nanostructure and morphology of materials.

Its new generation of SAXS system, the  Xeuss 2.0, combines small and wide-angle X-ray scattering techniques (SAXS/WAXS) in a proprietary technology which is helping the company lead the way to the ultimate laboratory nanostructure characterization tool.

The company also launched this year a new vertical SAXS/WAXS system called the Nano-inXider which comes in a compact package for integration in any lab environment.

Xenocs is operating worldwide through a strong network of distributors ensuring local support and close contact to customers. Hundreds of research groups around the world as well as many renowned Original Equipment Manufacturers of the X-ray analytical market trust Xenocs to provide them with the best solution to move ahead in their research activities and industrial applications.

Xenocs is a spin-off company from the Laue Langevin Institut in Grenoble, France.

This information has been sourced, reviewed and adapted from materials provided by Xenocs.

For more information on this source, please visit Xenocs.

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