P R O T E I N S I N D E V E L O P M E N T
243
lower than in the controls. On the other hand the rate of decrease in
viscosity of proteins treated with animalizing agents is much higher than
in the controls (Ranzi and Citterio, 1954). These data show the greater
resistance of proteins treated with vegetalizing substances and the lower
resistance of proteins treated with animalizing substances. The experiment was repeated with other substances. All the vegetalizing substances exert the same action of protecting from denaturation. All
animalizing agents denature and predispose to denaturation.
A similar experiment was carried out with a monodisperse globular
protein, the lipovitellin from hen eggs prepared according to Fujii
(1960) and the same results were observed. Because of the globular
shape of its molecules, viscosity increases with denaturation; consequently the rate of increase in viscosity was taken as an index of denaturation. The increase in viscosity after incubation with urea was less
in Li Cl-treated lipovitellin than in the control solution and greater in
NaSCN- or IBA-treated lipovitellin (Table 2).
TABLE 2
Resistance of lipovitellin to urea: Values =
where η ί is the specific viscosity
Vc
of the treated sample after incubation with urea and 77 c is the viscosity of the same
sample diluted with the solvent instead of urea
Treated with Li
Control
Treated with IBA
Treated with NaSCN
104·4±2·47
114·4±0·41
115-8
1181 ±0-81
The same type of experiment can be used to study the action of the
proteolytic enzyme, trypsin, on protein treated with LiCl or with animalizing substances. We prepared hen lipovitellin as before and treated
different samples with LiCl, NaSCN, or o-iodosobenzoic acid. The control
was represented by a solution of lipovitellin at the same concentration
in 2 M-NaCl. The samples, after very careful dialysis with 2 M-NaCl in
order to eliminate the substances added to the protein solution and to
obtain a concentration of NaCl equal to 2M in all the samples, were
digested with trypsin. Samples from each solution were taken every
three minutes and diluted 1 : 1 with trichloroacetic acid. The optical
density of the supernatant was read at 275 ηΐμ, in the Beckman spectrophotometer. The curves (Fig. 34) show that less protein is digested in the
solution treated with LiCl and more after treatment with NaSCN or IBA.
Consequently we may conclude that the denaturation induced by
animalizing substances leads to an additional denaturation induced by
urea; the action of vegetalizing substances is, however, a stabilizing
action that opposes the breakdown by urea. The same effect may be
243
lower than in the controls. On the other hand the rate of decrease in
viscosity of proteins treated with animalizing agents is much higher than
in the controls (Ranzi and Citterio, 1954). These data show the greater
resistance of proteins treated with vegetalizing substances and the lower
resistance of proteins treated with animalizing substances. The experiment was repeated with other substances. All the vegetalizing substances exert the same action of protecting from denaturation. All
animalizing agents denature and predispose to denaturation.
A similar experiment was carried out with a monodisperse globular
protein, the lipovitellin from hen eggs prepared according to Fujii
(1960) and the same results were observed. Because of the globular
shape of its molecules, viscosity increases with denaturation; consequently the rate of increase in viscosity was taken as an index of denaturation. The increase in viscosity after incubation with urea was less
in Li Cl-treated lipovitellin than in the control solution and greater in
NaSCN- or IBA-treated lipovitellin (Table 2).
TABLE 2
Resistance of lipovitellin to urea: Values =
where η ί is the specific viscosity
Vc
of the treated sample after incubation with urea and 77 c is the viscosity of the same
sample diluted with the solvent instead of urea
Treated with Li
Control
Treated with IBA
Treated with NaSCN
104·4±2·47
114·4±0·41
115-8
1181 ±0-81
The same type of experiment can be used to study the action of the
proteolytic enzyme, trypsin, on protein treated with LiCl or with animalizing substances. We prepared hen lipovitellin as before and treated
different samples with LiCl, NaSCN, or o-iodosobenzoic acid. The control
was represented by a solution of lipovitellin at the same concentration
in 2 M-NaCl. The samples, after very careful dialysis with 2 M-NaCl in
order to eliminate the substances added to the protein solution and to
obtain a concentration of NaCl equal to 2M in all the samples, were
digested with trypsin. Samples from each solution were taken every
three minutes and diluted 1 : 1 with trichloroacetic acid. The optical
density of the supernatant was read at 275 ηΐμ, in the Beckman spectrophotometer. The curves (Fig. 34) show that less protein is digested in the
solution treated with LiCl and more after treatment with NaSCN or IBA.
Consequently we may conclude that the denaturation induced by
animalizing substances leads to an additional denaturation induced by
urea; the action of vegetalizing substances is, however, a stabilizing
action that opposes the breakdown by urea. The same effect may be