The number of cooperative regions (energetic domains) in a pepsin molecule depends on the pH of the medium

Makarov, Alexander А.; Protasevich, I.I.; Frank, Ekaterina G.; Grishina, I.; Bolotina, I. A.; Ng, Esipova

doi:10.1016/0167-4838(91)90570-p

Cited by 18 publications

(19 citation statements)

References 13 publications

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“…1, ethanol addition has no effect on the shape of calorimetric curve, but results in the denaturation peak shift towards lower temperatures. The latter agrees with the results obtained for pepsin [6], ribonuclease A [1,2], lysozyme [3], cytochrome c [4] and ubiquitin [5]. It has been suggested that alcohols destabilize proteins by weakening hydrophobic interactions between non-polar residues as well as by disturbing the specific water structure around the protein molecule [3].…”

Section: Methodssupporting

confidence: 88%

“…The effect of ethanol on pepsin [6] and methanol on ribonuclease A [2] is similar. The AHc~ in both aqueous and water-ethanol solutions appears as a linearly increasing function of Td (within the studied pH range; Fig.…”

Section: Resultsmentioning

confidence: 81%

“…We have shown that 20% ethanol decreases the denaturation temperature of pepsin compared to that in aqueous solution, but has no effect on the calorimetric enthalpy of denaturation and the number of energetical domains (the regions melting according to the two-state model) [6]. This fact was observed only for neutral pH values at which the protein was not active.…”

Section: Introductionmentioning

confidence: 80%

“…This fact was observed only for neutral pH values at which the protein was not active. In 20% ethanol a pH decrease from 6.5 to 2.0 resulted in a decrease in the number of energetical domains in pepsin from four to two [6]. To determine the mechanism of ethanol effect on the thermal denaturation parameters at different pH values we have chosen to use pepsinogen, an inactive precursor of pepsin, differing from it by an additional positively charged 44-residue N-terminal prosegment.…”

Section: Introductionmentioning

confidence: 99%

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Heat denaturation of pepsinogen in a water‐ethanol mixture

et al. 1995

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The effect of ethanol and pH on thermodynamic parameters and cooperativity of pepsinogen heat denaturation was studied by scanning microcalorimetry. Addition of 20% ethanol decreases the protein denaturation temperature by 10.7°C at pH 6.4 and 15.8°C at pH 8.0. It also decreases the denaturation heat capacity increment from 5.8 to 4.2 kcal/K, mol. The dependences of calorimetric denaturation enthalpy on denaturation temperature both in water and 20% ethanol are linear and intersect at about 95°C. In 20% ethanol the pH shift from 5.9 to 8.0 results in a decreased number of cooperative domains in pepsinogen. This process causes no changes either in the secondary structure or in the local surroundings of aromatic amino acids. It is concluded that ethanol addition does not affect the cooperativity of pepsinogen denaturation substantially until the pH change provokes redistribution of charges in the protein molecule.

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Section: Methodssupporting

confidence: 88%

Section: Resultsmentioning

confidence: 81%

Section: Introductionmentioning

confidence: 80%

Section: Introductionmentioning

confidence: 99%

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Heat denaturation of pepsinogen in a water‐ethanol mixture

et al. 1995

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“…at the same denaturation temperature shows good coincic ence [16]. The ratio R = AH,JAHe~ is a criterion which allows ne to estimate how much the average size of a cooperative tnit, i.e., the region of a protein molecule which melts more or I~ss independently, differs from the size of the whole protein ~lobule [15,17]. T!…”

Section: Electrochemistrymentioning

confidence: 99%

Thermal stability of the polyheme cytochrome c₃ superfamily

et al. 1995

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Decoupling of melting domains in immobilized ribonuclease A

Rialdi

Battistel

1994

Proteins

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Ribonuclease A has been immobilized on silica beads through glutaraldeyde-mediated chemical coupling in order to improve the stability of the protein against thermal denaturation. The thermodynamic and binding properties of the immobilized enzyme have been studied and compared with those of the free enzyme. The parameters describing the binding of the inhibitor 3'-CMP (Ka and delta H) as monitored by spectrophotometry and calorimetry were not significantly affected after immobilization. Conversely both the stability and unfolding mechanism drastically changed. Thermodynamic analysis of the DSC data suggests that uncoupling of protein domains has occurred as a consequence of the immobilization. The two state approximation of the protein unfolding process is not longer valid for the immobilized RNase. Protein stability strongly depends on the hydrophobicity properties of the support surface as well as on the presence of the inhibitor and pH. For example, after immobilization on a highly hydrophobic surface, the enzyme is partially in the unfolded state. The binding of a ligand is able to reorganize the protein structure into a native-like conformation. The refolding rates are different for the two protein domains and vary as a function of pH and presence of the inhibitor 3'-CMP.

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The number of cooperative regions (energetic domains) in a pepsin molecule depends on the pH of the medium

Cited by 18 publications

References 13 publications

Heat denaturation of pepsinogen in a water‐ethanol mixture

Heat denaturation of pepsinogen in a water‐ethanol mixture

Thermal stability of the polyheme cytochrome c₃ superfamily

Decoupling of melting domains in immobilized ribonuclease A

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The number of cooperative regions (energetic domains) in a pepsin molecule depends on the pH of the medium

Cited by 18 publications

References 13 publications

Heat denaturation of pepsinogen in a water‐ethanol mixture

Heat denaturation of pepsinogen in a water‐ethanol mixture

Thermal stability of the polyheme cytochrome c3 superfamily

Decoupling of melting domains in immobilized ribonuclease A

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Thermal stability of the polyheme cytochrome c₃ superfamily