Hubert Bradford Vickery

The History of the Discovery of the Amino Acids II. A Review of Amino Acids Described Since 1931 as Components of Native Proteins

Publisher Summary In the 40 years that have elapsed since the publication of the review of the protein amino acids in 1931, a surprising number of new amino acids have been found in the hydrolyzates of proteins. Most of these have a rather restricted distribution and many of them appear to be the products of the interaction of one or other of the classical amino acids present in the already assembled amino acid chain. This chapter reviews the available information on the most important of these new amino acids. They fall into two categories, the first being the classical twenty, really twenty-one if cysteine and cystine are counted separately. However, cysteine is an amino acid incorporated into the peptide chain, the oxidative cross-linking to form cystine probably being a secondary reaction. The second category includes amino acids presumably derived from one or other of the twenty enzyme-directed reactions of already assembled polypeptide chains. However, much remains to be learned of such interactions.

EVIDENCE FROM ORGANIC CHEMISTRY REGARDING THE COMPOSITION OF PROTEIN MOLECULES

The study of proteins has undergone a very interesting sequence of changes when considered from the standpoint of the scientific background of the workers in the field. The earliest contributors were for the most part affiliated with medicine. Beccari (1682-1766), who in 1747 prepared the first protein of vegetable origin, was both physician and natural philosopher. At various stages in his career he taught chemistry, physics, mathematics, logic, anatomy, and medicine, and made definite contributions in the fields of human nutrition, of weather observation and of disease. Fourcroy (1755-1809), to whom we owe the terms albumin and gelatin, was a distinguished physician and organizer of medical education. Scheele (1742-1786), who carried out the first noteworthy chemical investigation of casein, was an apothecary. Braconnot (178&1854), who discovered glycine and leucine in 1820, was also a t one stage in his career a pharmacist. Wollaston (1766-1828), the discoverer of cystine, was a physician and physicist as well as chemist. From such investigators one would expect shrewd factual observation but not analysis nor theory and, even in the next stage, when organic chemists assumed the burden of investigation, attempts to account for observations were rare. Berzelius supplied nomenclature (cystine, glycine) ; Mulder coined the word protein and advanced the first speculation on what these substances were, thereby stimulating much research ; Liebig discovered tyrosine and taught Ritthausen, the first of the great protein chemists. The period from 1820 to 1900 was one in which organic chemists and those whose interest in physiology converted them into the first generation of biochemists, such men as Kiihne, Hofmeister, Kossel, and Schulze, advanced our knowledge enormously. They laid a foundation that made lthe emergence of a Fischer inevitable, and perhaps their greatest contribution was the discovery of the various amino acids of which the protein molecule is formed. Under the spur of Fischer’s formulation of an acceptable protein theory, as well as of his own discovery of no less than three protein amino acids, progress

Artificial Enrichment of Beet Root Tissue with Glutamine

Summary The glutamine content of beet root tissue to be used for the preparation of glutamine on the large scale can be materially increased if the roots of the plants with tops attached as purchased in the market are immersed in 0.15 M ammonium sulfate solution for from 4 to 6 days before being trimmed and extracted with water. Tissue that will yield 5 g or more of glutamine per kg can be prepared in this way. A demonstration is afforded that, contrary to earlier observations, significant losses of glutamine are experienced if beet root tissue is dried at 80°C in preparation for analysis.

THE CONTRIBUTION OF THE ANALYTICAL CHEMIST TO PROTEIN CHEMISTRY

The present conference on methods for the determination of the amino acids that are produced by the hydrolysis of proteins has, as its primary object, the discussion of analytical techniques that have been developed in recent years. As an introduction, it seems desirable to consider the reasons why the analytical chemistry of the amino acids has attracted so much attention in the past, and to attempt to assess the value to science of the information that has been provided. The point of view of those who have concerned themselves with the fundamental problem of the composition of the protein molecule has undergone marked changes with the passage of the years. The earliest analyses of proteins were carried out in order to characterize and differentiate protein preparations from various sources. Subsequently, there developed an interest in the composition of the protein molecule as a whole : in particular, the desire to see if it is composed exclusively of units that yield amino acids on hydrolysis, or if units of some other kind are involved to any significant extent. In recent years, although both of the original motives have maintained their place in the thoughts of protein chemists, there has arisen a conviction that not only the chemical properties of the proteins, but also the physical properties, or at least many of them, can be assigned a rational explanation in terms of the amino acid composition, if this is sufficiently well known and can be adequately interpreted. Finally, still far on the horizon, but becoming ever brighter, is the hope that we shall ultimately be able to formulate a comprehensive and convincing theory of the structure of the protein molecule, a single brief statement that will shed light alike upon the chemical. physical, enzymological, immunological, and, in appropriate cases, the hormonal properties of these substances, and indicate the way for all subsequent research upon them. It is my purpose to trace briefly the reasons for these changes in the ambition of the protein chemist, but the point that I wish to emphasize at the start is that, in spite of the devotion and zeal with which the many problems of protein analysis have been attacked for more than a century, we have not yet attained a final solution. In the papers that are to follow

Preparation of Gliadin and Zein.

Summary Methods are described for the preparation of gliadin from wheat and of zein from corn (maize). The products are of a grade suitable for many types of nutrition investigations.

Proportion of Cystine Yielded by Hemoglobins of the Horse, Dog, and Sheep.∗

The presence of cystine among the products of hydrolysis of horse hemoglobin has escaped the notice of most investigators although Abderhalden, 1 about 30 years ago, reported the isolation of 0.3% from this protein. We have recently applied the cysteine cuprous mercaptide method 2 to the analysis of horse hemoglobin and have found 0.41% of cystine to be yielded by a highly purified preparation; it therefore seemed of interest to investigate the hemoglobins of other species, particularly in view of the fact that differences in sulfur content of various hemoglobins have long been a matter of record. Hemoglobin was prepared from the blood of the sheep and the dog, and was recrystallized twice by the method of Hoppe-Seyler. Each preparation was dissolved in water and the solution was poured into a large volume of boiling water containing a trace of sodium chloride. The coagulated proteins were separately washed by repeated centrifugation with water until free from electrolytes, and were then dried in a current of warm air and ground to a fine powder. The yield of cystine after hydrolysis in the presence of tin was determined by the cuprous mercaptide method, 2 iron was determined by the method of Kennedy 3 with the modifications of McFarlane, 4 †sulfur was determined by the method of Denis 5 after a preliminary oxidation of the protein with hot concentrated nitric acid, and nitrogen was determined by the Kjeldahl method with mercury as catalyst. The results of these analyses are shown in the table where they are calculated on an ash- and moisture-free basis. The differences in the yields of cystine from these 3 hemoglobins are particularly striking and suggest that the yields of other amino acids from hemoglobins of different origin may likewise vary widely.

Potassium Dihydrogen D8L8-Isocitrate

The monopotassium salt is to be preferred to the lactone as the final product of the synthesis of isocitric acid by the method of Fittig and Miller.

Nutritive Properties of the Seed of the Tobacco Plant (Nicotiana tabacum)20

Although every part of the tobacco plant has been reported to contain nicotine, this alkaloid could not be detected by Vickery and Pucher in the fully ripened seed of Connecticut shade-grown tobacco by chemical methods. 1 Ilyin, 2 who has studied the distribution of nicotine in the plant, found that immature seed, and particularly the ovules at an early stage of development, contained small proportions, but that, as ripening progressed, the alkaloid content diminished until finally none could be demonstrated. In view of the toxic properties of nicotine it seemed that a simple physiological test for its presence in tobacco seed would consist in feeding trials on small animals. We therefore offered to albino rats a ration that consisted either of ground tobacco seed 98%, Osborne-Mendel salt mixture 3 2%, or ground tobacco seed 99%, sodium chloride 0.5%, calcium carbonate 0.5%; cod liver oil was administered as a supplement at the rate of 10 drops per day. The diet was consumed with avidity and without any evident untoward consequences; the animals grew at a satisfactory rate and appeared to be normal in every respect. This somewhat surprising outcome led to a detailed study of the nutritive properties of the tobacco seed.