Amino Acids, Peptides and Proteins - Dévényi T., Gergely J. 1976

Selected methodological aspects of analytical protein research
Study of native proteins

The rapid development of several branches of biology began only a century ago. At about the same time, the systematic study of Proteins was initiated, which is to some extent a more difficult task than, for example, solving certain problems in physiology or pharmacology. The primary difficulty in protein research lies in the fact that the subject of study is a very large and, apparently, highly labile molecule with an exceptionally complex Structure. Consequently, The Development of Methods for isolating and studying native, undenatured proteins proceeded rather slowly. However, over the past two decades, as the development of protein analysis methods entered a phase of rapid and comprehensive growth, Protein Chemistry has made a giant leap forward. Among the outstanding achievements of the last 10—15 years, the elucidation of the Introduction/19.html">Primary Structure of several proteins occupies a prominent place. At the same time, even today, studying The structure of any reasonably complex protein remains a highly challenging task.

In recent years, thanks to the development of Analytical Methods, the Study of native Proteins has become increasingly important in chemical, biochemical, and medical research, as well as in clinical practice. In a book aimed at providing a detailed description of protein analysis methods, it would be difficult to present a comprehensive review of the results obtained using each specific method; quite naturally, this should not be expected from a manual designed for specific practical tasks. We decided that it would be most appropriate to illustrate the capabilities and limitations of each analytical method using carefully selected Examples.

A good example of studying native proteins is the investigation of Blood Serum proteins, which includes their isolation, purification, and quantitative and qualitative analysis. The widespread application of protein analysis methods will be demonstrated precisely through The Study of serum proteins.

While opening up very broad Prospects for protein analysis in general, various electrophoretic and immunochemical Research Methods, as well as Chromatography and Gel filtration, have simultaneously enabled the discovery of several new serum proteins. This, in turn, not only helped clarify the details of certain physiological and pathophysiological processes, but also significantly stimulated the study of fundamental immunological phenomena, the elucidation of Cell/13.html">Protein Structure, etc. Of course, the material we present on serum proteins cannot be considered exhaustive. We aimed only to introduce the reader to the possibilities offered by each of the described methods, and we hope that this manual will prove useful in selecting the most appropriate method, as well as in interpreting the results obtained, for researchers working with both serum and other proteins.

In 1878, Hammarsten succeeded in separating globulins from serum albumins by salting out with saturated magnesium sulfate. This discovery dramatically changed the prevailing understanding of serum proteins at the time. The Use of magnesium sulfate initiated a whole series of protein Fractionation Methods based on salting out. For several decades, the salting-out Procedure was the only available method in both clinical and scientific studies of blood proteins. At this stage, serum proteins were characterized by their solubility in Water and in salt solutions of various concentrations. Based on these data, a distinction was made between the water-insoluble fraction, euglobulin, and the soluble fractions, pseudoglobulin and albumin.

The classical definition of serum globulins is associated with their salting out with ammonium sulfate, since globulins are defined as proteins that precipitate upon half-saturation of serum with this salt.

Protein fractionation by salting out is not without its drawbacks. It does not provide a sufficiently sharp Separation of the individual components of a mixture. The salt concentrations that precipitate a given protein fraction of normal serum may differ from those required for fractionating sera containing “pathological” proteins, etc. Despite this, salting out is still widely used in clinical laboratories to separate globulins and albumin, as this technique is a relatively simple and rapid METHOD FOR DETERMINING the albumin-globulin ratio (A/G ratio).