Accelerated X-ray structural analysis of proteins
The new Pepsi-SAXS protein analysis method works 50 times faster than analogues
Scientists at the University of Grenoble and MIPT, led by Sergey Grudinin, have created a highly efficient method for calculating X-ray scattering curves for analyzing protein molecules in Pepsi-SAXS solution, which works from 5 to 50 times faster than its analogues. The results are published by the International Union of Crystallography in the journal Acta Crystallographica (Grudinin et al., Pepsi-SAXS: an adaptive method for rapid and accurate computation of small-angle X-ray scattering profiles).
Proteins have a complex structure and an extremely small size – on the order of a few nanometers. To study them, it is necessary to invent unusual methods, since any impact can destroy the sample or change its properties. Knowledge about the structure of biomolecules and the mechanisms of their work allows us to develop new drugs not by trial and error (high-performance screening, strictly speaking), but based on a rational basis.
One of the ways of studying is the analysis of X-rays reflected from the protein. Scientists use X-ray light, not ordinary light, because they are interested in atomic sizes, about 1/10000 microns. Visible light is a little less than a micron. Therefore, in order to look at such small objects, light with a very short wavelength is needed. This property is exactly what X-rays have.
Maria Garkavenko, co-author of the paper, MIPT student: "The Pepsi-SAXS method allows efficient and highly accurate construction of scattering curves, as well as analysis of the three-dimensional structure of the sample. Eg, Pepsi-SAXS makes it possible to increase the efficiency of modeling and prediction of three-dimensional structures of macromolecules and much more."
Small–angle X-ray scattering (SAXS) is a type of small-angle scattering in which X-rays are scattered from a sample and then collected at very small angles. A graph of the dependence of the scattering intensity on the angle of incidence is obtained. Based on this graph, a comparative analysis is carried out with the experimental database of samples, and then a conclusion is made about the structure and properties of the protein under study.
Compared to other methods of structure determination, SAXS is much simpler and cheaper. No long special preparation of samples, freezing or crystallization of protein compounds is required. Samples are measured directly in the solution in a functional state. Thus, the reliability of the results is seriously improved, since during preparation the sample can change its state and properties. Another very important advantage of the method is that the destructive effect of X-rays on the experimental sample is insignificant.
But until recently, the SAXS methods had a significant drawback – the complexity of calculations, greatly limiting the number of experiments. It took about 10 hours to process the data of only one experiment. In the standard approach, the number of calculations is directly proportional to the square of the number of molecules of the sample, and this number was more than a thousand. The first idea to simplify calculations came to the German scientist Heinrich Navigator in the 70s of the last century. He proposed to describe the scattering from molecular compounds using special spherical functions. Despite the fact that the Navigator, due to the lack of computers, had to manually perform calculations on paper, even then the approach showed its effectiveness. A lot has been done in this area by graduates of the Soviet school, in particular, Dmitry Svergun (now working in Hamburg), who wrote a large Atsas package for all kinds of aspects of small-angle scattering on biomolecules. The researchers used these developments in their work.
Andrey Kazennov, co-author of the paper, graduate student at MIPT: "Pepsi-SAXS stands for "Polynomial expansions of protein structures and interactions' Small-angle X-ray Scattering" – an adaptive method for fast and accurate calculation of small-angle X-ray scattering profiles. Pepsi-SAXS can adjust to the size of the analyzed sample and the accuracy of experimental data."
Additionally, scientists have created an effective model of the water shell of the analyzed protein compounds, which seriously improves the accuracy of the results of the technique.
Sergey Grudinin , Head of the study: "The method was tested on a large sample of data collected from two of the largest biological databases, BioIsis and SASBDB. We have demonstrated that Pepsi-SAXS works from 5 to 50 times faster than the previously used methods of CRYSOL, FoXS and the three-dimensional Zernike method in SAStbx. At the same time, Pepsi-SAXS is not only not inferior to them in accuracy, but even wins."
In addition, special attention was paid to the analysis of the obtained results and their automatic comparison with experimental data.
Figure 1 shows the results of one of the series of experiments – a comparison of the work of various calculation methods used today on the SASDAW3 sample from the SASBDB database. The graph shows the average scattering intensity from the scattering angle. It is worth noting that the error of the calculated model χ2 has the lowest value in the Pepsi-SAXS model, which proves its high accuracy.
Fig. 1. Operation of the methods on the SASDAW3 sample
The study of protein compounds is of fundamental importance for understanding the processes of vital activity of living organisms, the creation of medicines and methods of treating diseases, as well as obtaining new organic materials up to the cultivation of artificial organs. With the new invention of our scientists, work in these areas will be able to occur 50 times faster.
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08.06.2017