hydrogen sulphide, exert a marked retarding influence on the
proteid-digesting power of trypsin. Moreover, while it is generally
stated that proteolytic and other enzymes are practically indifferent
to the presence of chloroform, thymol, and other like substances
that quickly interfere with the processes of the so-called organized
ferments, pepsin and trypsin certainly do show a certain degree of
sensitiveness to chloroform, and indeed even to a current of air
passed through their solutions. Thus, very recently, Bertels[40] and
Dubs[41], working under Salkowski’s direction, have called attention
to the peculiar behavior of pepsin to chloroform; their results
showing, first, that small amounts of this agent tend to increase the
proteolytic power of the enzyme, while larger amounts decrease its
digestive power. Another interesting point brought out especially
by Dub’s experiments is the fact that an impure solution of the
ferment, viz., an acid extract, for example, of the mucous membrane
of the stomach containing more or less albuminous matter, is far less
sensitive to chloroform than an acid solution of the purified ferment,
thus showing again the protective influence of proteids and other
extraneous matters; the latter guarding the enzyme to a certain extent
from both the stimulating and inhibitory action of various agents.
[40] Ueber den Einfluss des Chloroforms auf die Pepsinverdauung,
Virchow’s Archiv, Band 130, p. 497.
[41] Der Einfluss des Chloroforms auf die künstliche Pepsinverdauung,
Ibid., Band 134, p. 519.
Another point to be emphasized just here is that any chemical
substance, such as a metallic salt, having a specific action upon
proteid matter, will almost invariably interfere more or less with
the proteolytic action of these enzymes, both through a direct
action upon the soluble ferment itself, and also through an indirect
action in modifying or inhibiting the digestibility of the proteid
exposed to proteolysis. All of these facts emphasize more or less the
proteid-like nature of the enzymes, or at least the carriers of the
ferments. It is further very suggestive that the destruction of these
enzymes by heat happens to occur at approximately those temperatures
which are generally recognized as the coagulation points of ordinary
proteids. Moreover, the apparent lack of stability so characteristic of
these ferments, their inherent proneness to alteration, their marked
susceptibility to every change in environment, all point to large
complex molecules, such as we have in proteids and are familiar with in
living protoplasm.
Whatever the exact nature of these proteolytic enzymes, they are
certainly endowed with the power of transforming relatively large
amounts of proteid matter into soluble products, even though they
themselves are present in very small quantity. They are derived, as
we have seen, from living protoplasmic cells, and we might perhaps,
with v. Nägeli[42] and Mayer,[43] consider them as retaining a portion
of that molecular motion so characteristic of living protoplasm, by
which the equilibrium of the dead food-proteid may be disturbed and
thus changes started which result in what we call proteolysis. However
this may be, we must look to some phase of catalytic or contact
action as the true explanation of this power of proteolysis. At first
glance, any explanation or theory involving the use of catalysis seems
exceedingly vague and indefinite, and yet many illustrations can be
given of chemical reactions where the dominating agent evidently acts
in this manner. “We call a force catalytic,” says the philosopher of
Heilbron, “when it holds no communicable proportion to the assumed
results of its action. An avalanche is hurled into the valley....
A puff of wind or the fluttering of a bird’s wing is the catalytic
force which has given the signal for, and which is the cause of the
wide-spread disaster.”[44] In the older theories of catalytic action,
the catalytic agent was supposed to remain passive, but not so in the
more modern conception of catalysis. The ferment by its presence makes
possible certain changes and combinations which could not occur in its
absence, at least under the existing circumstances, although all the
other conditions might be favorable. The proteids, for example, have
a natural tendency to undergo hydration; thus, simple boiling with
dilute acid or exposure to the action of superheated water alone,[45]
will produce many if not all of the products formed in natural
digestive proteolysis. To be sure, they are not formed as readily as
in artificial or natural digestion, and there may be some minor points
of difference, but still proteolysis can be imitated in this manner.
The proteolytic enzymes simply help on this natural tendency of proteid
bodies to undergo hydration, and by their presence and action enable
it to occur at lower temperatures than it otherwise could, and at the
same time render it more rapid and complete. This is not accomplished,
however, by simple contact. The enzymes, we may assume, combine in
some manner with the proteid undergoing digestion, starting thereby a
train of reactions in which the proteid and the water present are the
main actors, they being, however, perfectly passive in the absence of
the inciting agent, the enzyme. As expressed by Ganger, “the ferment
phenomena resemble those in which there is apparently a periodic
synthesis and dissociation of the catalyzing agent, which acts in a
similar manner to the agent which explodes a train of gunpowder.”
[42] Theorie der Gährung. München, 1879.
[43] Die Lehre von den chem. Fermenten. Heidelberg, 1882.
[44] Quoted from Gamgee’s Physiological Chemistry of the Animal Body,
vol. 2, p. 7.
[45] Chittenden and Meara. A study of the primary products resulting
from the action of superheated water on coagulated egg-albumin. Journal
of Physiol., vol. 15, p. 501.
We can find many illustrations among chemical phenomena where one
body, even though present in small quantity, acts as a go-between and
makes possible an almost indefinite exchange of matter and energy.
Take as an illustration, the part played by water in determining the
explosion of oxygen and carbon-monoxide gas. Some years ago, Dixon[46]
called attention to the fact that a mixture of these two gases when
perfectly dry would not explode even by contact with red hot platinum
wire. The presence, however, of a small amount of aqueous vapor
would at once cause an explosion to occur. In confirmation of this
observation, Traube[47] has reported that a flame of carbon-monoxide
gas introduced into a perfectly dry atmosphere is at once extinguished.
In a moist atmosphere, on the other hand, the flame will continue to
burn indefinitely, that is as long as the CO gas is supplied. In these
cases, the water, which is so necessary for the appearance of the
reaction, and which furnishes a striking illustration of the action of
a contact or catalytic substance is not purely passive. To be sure,
only a minimal amount is necessary for the combustion of an indefinite
amount of carbonic oxide, but the water enters into the reaction
itself. It is to be noticed that carbonic oxide and water alone, even
at high temperatures, will not react, but in the presence of oxygen
the water is decomposed with formation of hydrogen-peroxide, thus:
CO + 2 H_{2}O + O_{2} = CO(OH)_{2} + H_{2}O_{2}.
[46] Chemical News, vol. 46, p. 151.
[47] Bericht. d. Deutsch. chem. Gesellsch., Band 18, p. 1890.
The hydrogen-peroxide thus formed combines with carbonic oxide to form
carbonic acid, which in turn is decomposed into the anhydride CO_{2},
with regeneration of water, the latter being available for further
action of the same order:
H_{2}O_{2} + CO = CO(OH)_{2}
2 CO(OH)_{2} = 2 CO_{2} + 2 H_{2}O.
Indeed, as can be readily seen from the equations, this may be kept
up indefinitely, a small amount of water, _i.e._, the go-between,
the catalytic agent, sufficing to accomplish the transformation of
almost any amount of carbon-monoxide. This, I think, furnishes an
excellent illustration of the way in which catalytic agents, such as
the proteolytic enzymes, may be supposed to act. It is truly contact
action, but the agent is not purely passive; the enzyme combines with
the substance undergoing proteolysis, and the resultant compound thus
formed is enabled now to combine with water and undergo hydrolysis,
something which could not be accomplished by the proteid and water
alone, that is at body temperature. This new and more complex compound
is naturally less stable and soon undergoes dissociation or cleavage
with a splitting off of the original enzyme for one product, which is
thus available for further action of the same order; while, as other
products, we find the hydrated and otherwise altered substances coming
from the proteid, and whose formation is the ultimate object of the
whole process.
The parallelism between this hypothetical action of the proteolytic
enzymes and the known reactions in the above combustion of carbonic
oxide is certainly very close, and leaves little doubt that this
explanation of enzyme action is, in a general way at least, correct.
Thus the carbonic oxide, CO, brought in contact with pure, dry oxygen
gas (apparently all that is necessary for its direct oxidation into
carbonic acid, CO_{2}), undergoes no change; the burning CO gas is at
once extinguished. Evidently, something more is necessary in order to
start the process of oxidation. So, too, in proteolysis; the process,
as we shall see later on, is essentially one of hydration, but bring
the proteid and the water, or acid-water, together and although all
the conditions are apparently favorable for hydration there is, as
you know, little or no change. But introduce the catalytic agent and
immediately the reaction commences. In the case of the burning CO
gas in contact with oxygen, the water acting as contact agent makes
oxidation possible, enabling the main actors in the transformation
to react upon each other. But, as we have seen, the contact agent is
something more than a mere looker-on, it becomes for the time being an
integral part of the molecule, undergoing change, combining with it and
thus making possible the subsequent alterations characteristic of the
specific transformation, in which, however, the regeneration of the
contact agent is a prominent feature. So, too, with the proteolytic
enzymes, pepsin and trypsin, they are the go-betweens, making possible
the union of the proteids with water by combining with the proteid
molecule and thus paving the way for both hydration and cleavage. In
the cleavage of the complex molecule, we have the regeneration of the
ferment as a prominent feature, and in proteolysis we understand that
the regenerated ferment may act not only upon more of the original
proteid, but likewise upon the primary products of its action, thus
giving rise eventually to a row of more or less closely related
cleavage products. Finally, we can conceive that the enzyme may
gradually be affected by the process, that its regeneration may become
less complete, and thus digestive power be eventually diminished.
Much more might be said in support of the above hypothesis. On the
other hand, some objections might be raised against it, but I know
of no more reasonable explanation of enzyme action than that here
presented, or one which so well accords with all of the known facts
concerning the conditions which modify proteolytic action.[48] Thus,
the influence of heat, of the products of proteolysis, of acids,
alkalies, and various organic and inorganic salts on the action of
these digestive enzymes is such as lends favor to the above view rather
than opposes it.
[48] Compare L. de Jager, Erklärungsversuch über die Wirkungsart
der ungeformten Fermente, Virchow’s Archiv, Band 121, p. 182. Also
Chandelon: Bulletin de l’Academie Royale de Méd. de Belgique, 1887, 1,
p. 289.
THE GENERAL NATURE OF PROTEIDS.
Proteids are confessedly among the most complex bodies the physiologist
has to deal with, while at the same time they are perhaps the most
important, not only in view of their wide-spread distribution through
animal and vegetable tissues, but because of the prominent part they
take in the nutrition of the body. The more our knowledge is broadened
concerning these varied substances, the more we are impressed with
their complexity, and at the same time with the necessity for a more
accurate study of both their composition and constitution. Concerning
the latter, full fruition of our hopes is probably in the distant
future, but every step of advance in this direction adds greatly
to our resources in the interpretation of the varied and complex
changes characteristic of proteid metabolism. Every study of proteid
decomposition adds something to our store of knowledge, and gives
perhaps an added fact available for broadening our deductions.[49]
Moreover, the composition and general reactions of the proteids
may be investigated with full confidence of obtaining many useful
results, which must necessarily be an aid in interpreting the changes
accompanying digestive proteolysis.
[49] See Drechsel: Der Abbau der Eiweisstoffe. Du Bois Reymond’s Archiv
f. Physiol., 1891, p. 248.
Take, for example, the single question of peptonization by gastric
digestion. What is the nature of the process? Is the proteid
transformed into a soluble and diffusible peptone as a result of
hydration and cleavage, or is it a transformation which results from
a simple depolymerization of the proteid molecule, _i.e._, are we to
consider albumin and peptone as isomeric bodies? These questions, on
which physiologists seem loath to agree, can certainly be answered
definitely; not, however, by arguments but by careful experimentation,
in which the composition of the proteid undergoing digestion must be
a necessary preliminary factor, and the composition of the resultant
product, or products, a secondary factor of equal importance. Further,
the question needs to be answered not with reference to one proteid
merely, but with reference to every proteid capable of digestion by
either gastric or pancreatic juice. When these questions have been
fully answered in this manner, we shall have positive data to deal
with, and our conclusions will rest upon a foundation of fact not
easily set aside. This is one of the problems upon which I have been at
work for some years, and although progress may in one sense be slow,
yet it is sure and gives results of no uncertain character.
First, then, let us consider briefly the nature of the proteids whose
proteolysis we may be interested in; remembering, however, that in so
doing we can merely touch upon the points essential for our purpose.
Allow me to say in parenthesis that there is being published in Moscow
a work on proteids alone of five volumes, 900 pages each, which it is
supposed will constitute an exhaustive treatise of the subject.[50]
[50] Die Einheit der Proteinstoffe. Historische u. experimentelle
Untersuchungen, by L. Morokhowetz.
If we attempt to classify all of the proteid bodies hitherto discovered
and studied we are at once confronted with a problem of no small
proportions. So varied are they in their reactions, solubilities, and
behavior toward general reagents, so inclined to merge into each other
by almost insensible gradations that it becomes an extremely difficult
matter to make an arrangement that will satisfy all the requirements
of the case. I have to suggest, however, the following classification,
which is merely a modification of several existing ones, based
primarily upon chemical composition, and solubility in the more common
menstruums.
Proteids may first be divided into three main groups as follows:
I. _Simple Proteids._--Composed of carbon, hydrogen, nitrogen, sulphur,
and oxygen, and yielding by decomposition aromatic bodies such as
tyrosin, phenol, indol, etc.
II. _Compound Proteids._--Composed of a simple proteid united to some
non-proteid body.
III. _Albuminoids._--A class of nitrogenous bodies related to and
derived from proteids, but differing especially from the latter by
great resistance to the ordinary solvents of true proteids.
The individual members of these three groups may be arranged as follows
on the basis of solubility, coagulability, etc.:
I. SIMPLE PROTEIDS.--_A._ _Soluble in water._--_a._ Coagulable by heat,
and by long contact with alcohol. Albumins: serum-albumin, egg-albumin,
lacto-albumin, myo-albumin, vegetable albumins. _b._ Non-coagulable by
heat and by long contact with alcohol. Proteoses:[51] protoproteoses,
deuteroproteoses. Peptones:[51] amphopeptones, antipeptones,
hemipeptones.
[51] Used in the generic sense; the proteoses including albumoses,
globuloses, myosinoses, elastoses, etc., and the peptones the
peptone-products formed in the digestion of any or all proteids.
_B._ _Insoluble in water, but soluble in salt solutions._--_a._ More or
less coagulable by heat. Globulins. 1. Soluble in dilute and saturated
NaCl solutions. Vitellins. 2. Soluble in dilute NaCl solutions, but
precipitated by saturation with NaCl. Myosins, paraglobulin[52]
or serum-globulin, fibrinogen, myo-globulin, paramyosinogen,
cell-globulins. _b._ Non-coagulable by heat, soluble in dilute NaCl
solution and precipitated by saturation with NaCl. Heteroproteoses.
[52] Not completely insoluble in saturated NaCl solution.
_C._ _Insoluble in water and salt solutions, soluble in dilute
alcohol_--Zein, gliadins.
_D._ _Insoluble in water, salt solutions and alcohol; soluble in
dilute acids or alkalies._--_a._ Coagulable by heat when suspended in
a neutral fluid. Acid-albumins, alkali-albumins or albuminates. _b._
Non-coagulable by heat when suspended in a neutral fluid. Antialbumids,
dysproteoses, glutenins.
_E._ _Insoluble in water, salt solutions, alcohol, dilute acids and
alkalies; soluble in strong acids, alkalies, and in pepsin-hydrochloric
acid and alkaline solutions of trypsin._--Coagulated proteids,
fibrin.[53]
[53] In blood-fibrin we have a good illustration of the fact that these
divisions are not absolutely exact, since this form of proteid matter,
for example, is somewhat soluble in dilute acids and in salt solutions,
although requiring a long time for marked solution.
II. COMPOUND PROTEIDS.--_A._ _Compounds of a proteid (globulin) with an
iron-containing pigment, soluble in water and coagulable by heat and
alcohol._ Hæmoglobin, oxyhæmoglobin, methæmoglobin, etc.
_B._ _Compounds of proteids with members of the carbohydrate group.
Insoluble in water; soluble in very weak alkalies._--_a._ True mucins.
_b._ Mucoids or mucinoids.
_C._ _Compounds of proteids with nucleic acid. Phosphorized bodies
yielding by decomposition metaphosphoric acid. Insoluble in water
and in pepsin-hydrochloric acid, but more or less soluble in
alkalies._--Nucleins.
_D._ _Compounds of proteids with nucleins. Very soluble in dilute
alkalies._--Nucleoalbumins, as casein of milk, and nucleoalbumins of
cell-protoplasm and cell-nuclei, etc.
III. ALBUMINOIDS.--_A._ _Soluble in boiling water with formation of
gelatin and yielding by decomposition leucin and glycocoll._--Collagen
(gelatin).
_B._ _Insoluble in boiling water, and yielding by decomposition
much leucin and some tyrosin, together with glycocoll and lysatin.
Slowly hydrated by boiling dilute acids and by treatment with
pepsin-hydrochloric acid._--Elastin.
_C._ _Insoluble in water, dilute acids and alkalies, also in
gastric and pancreatic juice. Yield leucin and tyrosin by
decomposition._--Keratin, neurokeratin.
We may now advantageously consider the composition of a few of the
more prominent representatives of the individual groups, taking
for illustration those bodies which have been most thoroughly
studied, and which we may have occasion to refer to in our
discussion of proteolysis. I have not included in the table any of
the alteration-products of the proteids formed by the action of
pepsin-acid, trypsin, or boiling dilute acids, confining myself here
simply to those bodies which occur ready-formed in nature.
COMPOSITION OF SOME OF THE MORE PROMINENT PROTEIDS OCCURRING IN
NATURE.[A]