Alkaline denaturation and partial refolding of pepsin investigated with DAPI as an extrinsic probe

Favilla, Roberto; Parisoli, Alessandra; Mazzini, Alberto

doi:10.1016/s0301-4622(97)00016-1

Cited by 28 publications

(30 citation statements)

References 17 publications

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“…[22,29] It has optimum activity in the pH range 1-4, minimal activity above pH 5 and is denatured at pH 6-7. [30,31] The single molecular chain of porcine pepsin (EC 3.4.23.1), which consists of 326 residues (34 623 kDa), adopts a bi-lobe structure composed of two topologically similar domains. [8,15,16] A graphical representation of pepsin is shown in Figure 1.…”

Section: Methodsmentioning

confidence: 99%

On the Compatibility Criteria for Protein Encapsulation inside Mesoporous Materials

et al. 2010

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The properties of the enzyme pepsin, relevant to its incorporation inside the channels of mesoporous silica materials in the preparation of bioinorganic hybrids, are highlighted by molecular dynamics simulations of aqueous solutions of the protein under conditions optimal for encapsulation in SBA-15. The protein size, shape, flexibility and surface properties are calculated with the aim of deriving general accessibility/compatibility criteria favouring encapsulation inside mesoporous systems.

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Section: Methodsmentioning

confidence: 99%

On the Compatibility Criteria for Protein Encapsulation inside Mesoporous Materials

et al. 2010

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“…21,22 However, a major consequence is that protein stability is often seriously compromised at neutral pH. [17][18][19][20] This is illustrated in the schematic energy landscape diagrams in Figure 8(c), where trypsin (stable at neutral pH) and pepsin (stable at low pH) are contrasted at both low and neutral pH.…”

Section: Acid Resistance Via Kinetic Stabilitymentioning

confidence: 99%

“…While this strategy increases stability at low pH by reducing positive electrostatic repulsion, there is a large increase in negative electrostatic repulsion and corresponding reduction in stability at neutral and alkaline pH. [17][18][19][20] Additional strategies include reducing the overall number of charged residues as well as the solvent accessibility of acidic residues (as in acidophilic maltose binding protein from Alicyclobacillus acidocaldarius 21 and N5-carboxyaminoimidazole ribonucleotide mutase from Acetobacter aceti 22 ) and the binding of metal or cofactors (as in rusticyanin from Thiobacillus. ferrooxidans 23,24 ).…”

Section: Introductionmentioning

confidence: 99%

Structural and Mechanistic Exploration of Acid Resistance: Kinetic Stability Facilitates Evolution of Extremophilic Behavior

Kelch

Eagen

Erciyas

et al. 2007

Journal of Molecular Biology

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“…Above pH 7, pepsin is in a denatured conformation that retains some secondary structure [14,15]. This denaturation is not fully reversible [16], the lack of reversibility being attributed to the N‐terminal domain [17]. In the 5–7 pH interval the conformation of pepsin is poorly characterised.…”

Section: Introductionmentioning

confidence: 99%

The active site of pepsin is formed in the intermediate conformation dominant at mildly acidic pH

Campos

Sancho

2003

FEBS Letters

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Pepsin is an aspartic protease that acts in food digestion in the mammal stomach. An optimal pH of around 2 allows pepsin to operate in its natural acidic environment, while at neutral pH the protein is denatured. Although the pH dependence of pepsin activity has been widely investigated since the 40s, a renewed interest in this protein has been fuelled by its homology to the HIV and other aspartic proteases. Recently, an inactive pepsin conformation has been identi¢ed that accumulates at mildly acidic pH, whose structure and properties are largely unknown. In this paper, we analyse the conformation of pepsin at di¡erent pHs by a combination of spectroscopic techniques, and obtain a detailed characterisation of the intermediate. Our analysis indicates that it is the dominant conformation from pH 4 to 6.5. Interestingly, its near UV circular dichroism spectrum is identical to that of the native conformation that appears at lower pH values. In addition, we show that the intermediate binds the active site inhibitor pepstatin with a strength similar to that of the native conformation. Pepsin thus adopts, in the 6.5^4.0 pH interval, a native-like although catalytically inactive conformation. The possible role of this intermediate during pepsin transportation to the stomach lumen is discussed.

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Alkaline denaturation and partial refolding of pepsin investigated with DAPI as an extrinsic probe

Cited by 28 publications

References 17 publications

On the Compatibility Criteria for Protein Encapsulation inside Mesoporous Materials

On the Compatibility Criteria for Protein Encapsulation inside Mesoporous Materials

Structural and Mechanistic Exploration of Acid Resistance: Kinetic Stability Facilitates Evolution of Extremophilic Behavior

The active site of pepsin is formed in the intermediate conformation dominant at mildly acidic pH

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