Molecular Biology: Protein. Enzymes and Coenzymes

The pattern of metabolism, sketched out in finer and finer detail as the mid-twentieth century passed, was, in a way, an expression of the enzymatic makeup of the cell. Each metabolic reaction is catalyzed by a particular enzyme and the nature of the pattern is determined by the nature and concentration of the enzymes present. To understand metabolism, therefore, it was desirable to understand enzymes.

Harden, who had begun the twentieth-century unravelment of intermediary metabolism, also unfolded a new aspect of enzymes. In 1904, he placed an extract of yeast inside a bag made of a semipermeable membrane (one through which small molecules might pass but not large ones) and placed it in water. The small molecules in the extract passed through and, after a while, the yeast extract could no longer break down sugar.

This could not be because the enzyme itself had passed through, since the water outside the bag could not break down sugar either. However, if the water outside were added to the extract inside, the mixture could break down sugar. The conclusion was that an enzyme (itself a large molecule unable to pass through a membrane) might yet include a relatively small molecule, loosely bound and therefore capable of breaking free and passing through the membrane, as part of its structure and essential to its function. The small, loosely bound portion came to be called a "coenzyme."

The structure of Harden's coenzyme was worked out, during the 1920s, by the German-Swedish chemist, Hans Karl von Euler-Chelpin (1873- ). Other enzymes were found to include coenzyme portions and the structure of a number of these was elucidated during the 1930s. As the molecular structure of vitamins was also determined in that decade, it became quite apparent that many of the coenzymes contained vitaminlike structures as part of their molecules.

Apparently, then, vitamins represented those portions of coenzymes which the body could not manufacture for itself and which, therefore, had to be present, intact, in the diet. Without the vitamins, the coenzymes could not be formed; without the coenzymes, certain enzymes were ineffective and the metabolic pattern was badly upset. The result was a vitamin-deficiency disease and, eventually, death.

Since enzymes are catalysts, needed by the body only in small quantities, coenzymes (and vitamins, too) are needed in small quantities only. This explains why a dietary component, present only in traces, may yet be essential to life. It was easy to see that minerals needed in traces, such as copper, cobalt, molybdenum, and zinc, must also form essential parts of an enzymatic structure, and enzymes containing one or more atoms of such elements have indeed been isolated.

But what of the enzyme itself? Throughout the nineteenth century, it had been a mysterious entity, visible only through its effects. The German-American chemist, Leonor Michaelis (1875-1949), brought it down to earth in a way by treating it according to physical-chemical principles. He applied the rules of chemical kinetics (a branch of physical chemistry that deals with the rates of reactions) and, in 1913, was able to derive an equation that described the manner in which the rate of an enzymecatalyzed reaction varies under certain set circumstances. To work out this equation, he postulated an intermediate combination of the enzyme and the substance whose reaction it catalyzed. This sort of treatment emphasized that enzymes were molecules that obeyed the physical-chemical laws to which other molecules were subject.

But what kind of a molecule was it? To be sure, it was strongly suspected of being a protein, for an enzyme solution easily lost its activity through gentle heating and only protein molecules were known to be so fragile. This, however, was only supposed and not proven, and during the 1920s, the German chemist, Richard Willstatter (1872-1942), advanced reasons for believing that enzymes were not proteins. His reasoning, as it turned out, was fallacious, but his prestige was great enough to lend his opinion considerable weight.

In 1926, however, the possibility that enzymes were proteins was raised again by an American biochemist, James Batchellor Sumner (1887-1955). In that year, Sumner was extracting the enzyme content of jack beans, the enzyme involved being "urease," one which catalyzed the breakdown of urea to ammonia and carbon dioxide.

In performing his extraction, Sumner found that at one point he obtained a number of tiny crystals. He isolated the crystals, dissolved them, and found he had a solution with concentrated urease activity. Try as he might, he could not separate the enzyme activity from the crystals. The crystals were the enzyme and all his tests further agreed on the fact that the crystals were also protein. Urease, in short, was the first enzyme ever to be prepared in crystalline form, and the first enzyme to be shown, incontrovertibly, to be a protein.

If further confirmation was wanting, or if the rule was suspected to be not general, the work of the American biochemist, John Howard Northrop (1891- ), finished matters. In 1930, he crystallized pepsin, the proteinsplitting enzyme in gastric juice; in 1932, he crystallized trypsin and, in 1935, chymotrypsin, both protein-splitting enzymes from pancreatic juice. These proved to be protein, too. Since then, dozens of enzymes have been crystallized and all have proved to be proteins.

By the mid-1930s then, the problem of enzymes had clearly merged with the general problem of proteins.