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Pressure acceleration of proteolysis: A general mechanism

View Article: PubMed Central - PubMed

ABSTRACT

Remarkable acceleration of enzymatic proteolysis by pressure at kbar range is reported with ubiquitin as substrate and α-chymotrypsin as enzyme. The acceleration is interpreted in terms of the shift of conformational equilibrium in ubiquitin from the non-degradable folded conformer to the enzyme-degradable unfolded conformer by pressure because of the lower volume of the latter, while the enzymatic activity of α-chymotrypsin is still largely retained. This mechanism is considered generally applicable to most globular proteins and the method of pressure-accelerated proteolysis will have an enormous potential utility in systems wherever efficient removal of proteins is needed.

No MeSH data available.


General mechanism for pressure enhancement of enzymatic proteolysis of a globular protein. Pressure shifts the population from N to U according to eq. 1 and U is subsequently degraded by an proteolytic enzyme, e.g. α-chymotrypsin under pressure.
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f3-4_29: General mechanism for pressure enhancement of enzymatic proteolysis of a globular protein. Pressure shifts the population from N to U according to eq. 1 and U is subsequently degraded by an proteolytic enzyme, e.g. α-chymotrypsin under pressure.

Mentions: For ubiquitin at pH 4.5 and 0°C, the stability at 1 bar (ΔGU-N0) was determined previously to be 31.3 +/− 4.7 kJ/mol, while the change in partial molar volume ΔVU-N to be −85 ml/mol at pH 4.5 and 0°C11. With an assumption of constant ΔVU-N over the range of pressure in eq. 1, we estimate that at 1 bar the population of U is less than 0.001%, which will be increased by an order of magnitude to ∼0.01% at 0.7 kbar, to ∼0.1% at 1.4 kbar, and to >1% at >2.0 kbar. Our proteolysis reaction had to be carried out at different conditions (pH 7.0, 37°C) and therefore the estimate of population U could be somewhat different from the above. Nevertheless, the above estimate demonstrates the fact that pressure within a few kbar range increases the population of the proteolysis-prone unfolded conformer dramatically, while a proteolytic enzyme, e.g., α-chymotrypsin, is still active (cf. Fig. 2). Thus, we propose a simple mechanism for the pressure enhancement of enzymatic proteolysis for a globular protein as shown schematically in Figure 3. Here the essence is that pressure shifts the conformational equilibrium between N and U in favor of U according to eq. 1, which is subsequently degraded by α-chymotrypsin under pressure.


Pressure acceleration of proteolysis: A general mechanism
General mechanism for pressure enhancement of enzymatic proteolysis of a globular protein. Pressure shifts the population from N to U according to eq. 1 and U is subsequently degraded by an proteolytic enzyme, e.g. α-chymotrypsin under pressure.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5036607&req=5

f3-4_29: General mechanism for pressure enhancement of enzymatic proteolysis of a globular protein. Pressure shifts the population from N to U according to eq. 1 and U is subsequently degraded by an proteolytic enzyme, e.g. α-chymotrypsin under pressure.
Mentions: For ubiquitin at pH 4.5 and 0°C, the stability at 1 bar (ΔGU-N0) was determined previously to be 31.3 +/− 4.7 kJ/mol, while the change in partial molar volume ΔVU-N to be −85 ml/mol at pH 4.5 and 0°C11. With an assumption of constant ΔVU-N over the range of pressure in eq. 1, we estimate that at 1 bar the population of U is less than 0.001%, which will be increased by an order of magnitude to ∼0.01% at 0.7 kbar, to ∼0.1% at 1.4 kbar, and to >1% at >2.0 kbar. Our proteolysis reaction had to be carried out at different conditions (pH 7.0, 37°C) and therefore the estimate of population U could be somewhat different from the above. Nevertheless, the above estimate demonstrates the fact that pressure within a few kbar range increases the population of the proteolysis-prone unfolded conformer dramatically, while a proteolytic enzyme, e.g., α-chymotrypsin, is still active (cf. Fig. 2). Thus, we propose a simple mechanism for the pressure enhancement of enzymatic proteolysis for a globular protein as shown schematically in Figure 3. Here the essence is that pressure shifts the conformational equilibrium between N and U in favor of U according to eq. 1, which is subsequently degraded by α-chymotrypsin under pressure.

View Article: PubMed Central - PubMed

ABSTRACT

Remarkable acceleration of enzymatic proteolysis by pressure at kbar range is reported with ubiquitin as substrate and α-chymotrypsin as enzyme. The acceleration is interpreted in terms of the shift of conformational equilibrium in ubiquitin from the non-degradable folded conformer to the enzyme-degradable unfolded conformer by pressure because of the lower volume of the latter, while the enzymatic activity of α-chymotrypsin is still largely retained. This mechanism is considered generally applicable to most globular proteins and the method of pressure-accelerated proteolysis will have an enormous potential utility in systems wherever efficient removal of proteins is needed.

No MeSH data available.