The mechanism by which an enzyme accomplishes its catalytic effects has been the object of extensive studies during recent years, especially since the discovery by Büchner that enzymes could be isolated in solutions entirely free from the disturbing influence of growing cells. Several theories concerning the mode of this catalytic action have been advanced. The earliest and simplest of these was that the enzyme simply creates an environment favorable for the particular chemical reaction to take place, as by exposing large surfaces of the substance in question to the action of the hydrolytic, or other effective, agent, by means of surface adsorption of the substrate material on the colloidal enzyme.
However, more recent investigations clearly indicate that there is an actual combination between the substrate material and the enzyme, which combination then breaks down with a resultant change in the substrate material and a freeing of the enzyme for repeated recombination with additional substrate, with the net result that the chemical change in the substrate material is enormously accelerated. That such a combination between substrate and enzyme actually exists has been demonstrated in two different ways: (a) experimentally, by mixing together solutions of an enzyme and of its substrate, each of which is filterable through paper or through a porous clay filter, with the result that the active material in the combined solutions will not pass through these same filters; and (b) mathematically, by a study of the curves representing the reaction velocities of typical reactions which are proceeding under the influence of an enzyme, which show that so long as there is a large excess of substrate material present, the accelerating influence of the catalyst is uniform over given successive periods of time, but that when the quantity of substrate material becomes smaller than that which permits the maximum combining power of the enzyme to be exercised, the reaction velocity immediately slows up.
Again, the fact that the specificity of the action of an enzyme, i.e., the limitation of the action of that enzyme to a specific single compound or group of similar compounds, is definitely related to the molecular configuration of the molecule of the substrate, as has been found to be true in all those cases where the molecular configuration of the substrate material has been established (see pages 56 to 58), is an added indication that there is some kind of a union between the enzyme and the substrate as a first step in the catalytic process.
As to the nature of this supposed combination of substrate and enzyme, two theories are held. The first is that this union is in the form of an actual molecular combination, or chemical compound, and the other is that it is a purely physical, or colloidal complex. The latter view has by far the greater weight of theoretical and experimental evidence in its support. The relation of electrolytes to the catalytic effect of enzymes, the appearance of the reacting masses under the ultramicroscope, and the effect of heat upon the reacting mixtures, all point to the conclusion that the phenomenon is colloidal rather than molecular in character. This view also makes the remarkable catalytic effects which take place in living protoplasm, which undoubtedly exists in the colloidal condition, much more easily understood. This phase of the matter will be much more apparent after the chapter dealing with the physical chemistry of the protoplasm has been studied.
A further indication that the mechanism of enzyme activity is colloidal in character lies in the fact that, so far as is known, all reactions which are catalyzed by specific enzymes are reversible and the same enzyme will accelerate the velocity of the reaction in either direction, the direction in which the reaction goes being determined by the conditions surrounding the reacting material at the time. It was formerly supposed that enzymes catalyze only decomposition reactions and that the synthetic reactions of living tissues are produced by means of some other force or agency. This view supported the idea of a chemical union of the enzyme with the substrate which, when it breaks down, breaks the molecule of the substrate material into some simpler form, or forms. But it is now known that the reaction which is influenced by the enzyme will be catalyzed in either direction by the specific enzyme which "fits" the particular substrate material at every point of its molecular configuration, as the glove fits the hand. The contrast between this fitting of the enzyme to the entire configuration of the molecule, and the union at a single point or group which is characteristic of chemical linkages, is apparent. As examples of the synthetic action of the same enzyme which, under other conditions, accelerates the decomposition of the same material, there may be cited the demonstrated synthesis of isomaltose from glucose by maltase; the production of ethyl butyrate from alcohol and butyric acid; and the synthetic production of artificial fats, by the aid of the pancreatic lipase; and the apparent synthesis of a protein from the same amino-acids which may be obtained from it by hydrolysis under the influence of the same protease, but under different environmental conditions.
ACTIVATORS AND INHIBITORS
The activity of enzymes is strongly influenced by the presence in the solution of other bodies, usually, although not always, electrolytes. This is probably due, in most cases at least, to the action of the electrolyte upon the colloidal condition of the enzyme. All enzymes do not respond alike to the action of the same electrolyte, however. The activity of certain enzymes is enormously increased by the presence of a small amount of acid; while the action of another may be absolutely inhibited by the same acid in the same concentration. Thus, the activity of the amylase found in the endosperm of many seeds is instantly stopped by adding to the solution enough sulfuric acid to make it two-hundredth normal in strength; while the same concentration of acid actually accelerates the activity of some of the proteases.
Formaldehyde, hydrocyanic acid, and soluble fluorides usually inhibit both the activity of a cell and of the enzymes which it contains; while other antiseptics, such as toluene, xylene, etc., prevent the growth of the cell, or organism, without interfering with the activity of the enzymes which may be present. By the use of this latter type of antiseptics, it is possible to distinguish between chemical changes which are involved in the actual development of a cell and those which can be brought about in other media by means of the enzymes which are contained in the cell.
Any substance which increases the catalytic activity of an enzyme is known as an "accelerator," or "activator"; while one which prevents this activity is called an "inhibitor," or "paralyzer."
A type of accelerating influence quite different from that of electrolytes is found in the effect of certain amino-acids upon enzyme action. The influence of small amounts of asparagine in enormously increasing the hydrolytic effect of amylase is an example. There is no known explanation for this type of activation of the enzyme.