Estimation of the stability of globular proteins

Ahmad, Faizan; Bigelow, Charles C.

doi:10.1002/bip.360250906

Cited by 44 publications

(30 citation statements)

References 26 publications

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“…This roughly compares with 0.634 obtained by Brandts (29) for the parameter p in his free energy expression for chymotrypsinogen. The surface area increases roughly by a factor of two upon unfolding, and this is consistent with earlier estimates made from entirely different experimental data (33,34). The area of the denatured protein is no more than about two-thirds that of the fully extended form.…”

Section: The Small Molecule Transfer Processessupporting

confidence: 90%

Isoenthalpic and isoentropic temperatures and the thermodynamics of protein denaturation.

Lee

1991

Proc. Natl. Acad. Sci. U.S.A.

116

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The standard enthalpy or entropy change upon transfer of a small nonpolar molecule from a nonaqueous phase into water at a given temperature is generally different for different solute species. However, if the heat capacity change is independent of temperature, there exists a temperature at which the enthalpy or the entropy change becomes the same for all solute species within a given class. Similarly, the enthalpy or the entropy change of protein denaturation, when extrapolated to high temperature assuming a temperatureindependent heat capacity change, shows a temperature at which its value becomes the same for many different globular proteins on a per weight basis. It is shown that the existence of these temperatures can be explained from a common formalism based on a linear relationship between the thermodynamic quantity and a temperature-independent molecular property that characterizes the solute or the protein. For the small nonpolar molecule transfer processes, this property is the surface area or the number of groups that are brought in contact with water. For protein denaturation, it is suggested that this property measures the polar/nonpolar mix of the internal interaction within the protein interior. Under a certain set of assumptions, this model leads to the conclusion that the nonpolar and the polar groups of the protein contribute roughly equally to the stability of the folded state of the molecule and that the solvent-accessible surface area of the denatured form of a protein is no more than about two-thirds that of the fully extended form.

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Section: The Small Molecule Transfer Processessupporting

confidence: 90%

Isoenthalpic and isoentropic temperatures and the thermodynamics of protein denaturation.

Lee

1991

Proc. Natl. Acad. Sci. U.S.A.

116

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“…3), which gives a midpoint for Gdn-HCI unfolding (C,,,) of 1.55 M and a corresponding free energy of stabilization of -6.8 kcal/mol (Table 1) (Hurle et al, 1994). These values are typical of globular proteins in this size range (Pfeil, 1981) and are also similar to values obtained for other V, domains (Ahmad & Bigelow, 1986;Tsunenaga et al, 1987).…”

Section: Resultssupporting

confidence: 72%

“…Studies on wild-type VL domains released by limited proteolysis of Bence-Jones proteins or expressed by recombinant DNA methods show that these immunoglobulin domains are about as stable as expected for proteins of about 100 amino acids, with AGsqab values around 3-9 kcal/mol (Ahmad & Bigelow, 1986;Tsunenaga et al, 1987;Hurle et al, 1994;Steipe et al, 1994;Stevens et a!., 1995). Wildtype REI V, falls within this range, with a AG:,, of about 6.5 kcal/mol (Hurle et al, 1994;Kolmar et al, 1994).…”

Section: Stability Of the Immunoglobulin Foldmentioning

confidence: 96%

Destabilizing loop swaps in the CDRs of an immunoglobulin V_L domain

Helms

Wetzel

1995

Protein Science

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It is generally believed that loop regions in globular proteins, and particularly hypervariable loops in immunoglobulins, can accommodate a wide variety of sequence changes without jeopardizing protein structure or stability. We show here, however, that novel sequences introduced within complementarity determining regions (CDRs) 1 and 3 of the immunoglobulin variable domain REI VL can significantly diminish the stability of the native state of this protein. Besides their implications for the general role of loops in the stability of globular proteins, these results suggest previously unrecognized stability constraints on the variability of CDRs that may impact efforts to engineer new and improved activities into antibodies.Keywords: complementarity determining regions; hypervariable loops; immunoglobulin stability; protein stability; Stern-Volmer plots; unfolding In the structures of antibody heavy and light chain variable domains, complementarity determining region (CDR) loops of diverse sequence and conformation project from P-sandwich frameworks to determine the affinity and specificity of antigen binding. These diverse sequences arise in the development of an antibody response via a complex combination of gene rearrangements and somatic mutations (Branden & Tooze, 1991). Protein engineers have transferred antigen binding activity from one framework to another by swapping hypervariable CDR loops between frameworks (Jones et al., 1986;Riechmann et al., 1988). In addition, totally artificial loop sequences have been introduced into variable domain frameworks to generate novel binding functions (Barbas et al., 1993;Fisch et al., 1994). This apparent tolerance of the immunoglobulin fold for both sequence diversity and loop swaps is consistent with the view that the major determinants of structural stability of globular proteins lie in the sequence-specific formation and packing of regular secondary structure elements (Shortle, 1992;Matthews, 1993), whereas surface loops are more forgiving of sequence changes (EL Hawrani et al., 1994).Although the CDR loops differ greatly in sequence, they are not infinitely variable in either sequence or conformation. Positions with highly conserved amino acids are found within many CDR sequences (Kabat et al., 1991 of "canonical structures" appear to be available for each CDR. For example, in an examination of the structural database available in 1989, only four different loop conformations were identified in the light chain crystallographic database for CDRl, and only three for CDR3 . If these patterns d o suggest constraints on the sequence variability of CDRs, however, it remains difficult from examination of evolved antibodies to either gauge the severity of these constraints or to understand their structural or functional basis.In this report we describe the preparation of a number of mutants of the immunoglobulin light chain variable domain REI, in which wild-type CDR sequences were replaced by loops of differing sequence and length. These replacements cause dramat...

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“…We were also interested in evaluating the sensitivity of protein stability to microenvironmental changes attained by the addition of those components. Although the concept of structural stability is unclear (Ahmad and Bigelow, 1986;Becktel and Schellman, 1987), a number of properties related to the conformational state of the protein, and therefore varying during unfolding, can be measured to monitor the conformational changes induced by heating (Robson and Gamier, 1986;Schmid, 1989). We used the melting temperature, namely, the temperature at which the concentrations of the native and denatured conformations are equal, as an index of stability.…”

Section: Introductionmentioning

confidence: 99%

Molecular thermodynamics of heat‐induced protein unfolding in aqueous media

Cioci

Lavecchia

1997

AIChE Journal

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The molecular thermodynamic model studied is based on the two-state mechanism of inactivation, in which only native folded and polymorphous unfolded protein forms are present at equilibrium. The influence of solvent on protein stability is described in terms of perturbation of the protein distribution between the two conformational states. An expression derived for the chemical potential of the protein accounts for conformational changes, ideal mixing effects, and interaction of the protein with the surrounding medium. Thermal unfolding of lysozyme was then studied in the absence or presence of hydroxylic compounds. Ultraviolet difference spectroscopy was used to monitor the conformational changes induced by heating and to determine the melting temperature of the protein. The additives investigated are ethanol, glycols, and natural osmolytes. Media containing ethanol and glycols destabilized lysozyme, whereas sugars increased the conformational stability of the protein. For all of the systems examined the melting temperature was linearly related to the surface tension of the mired solvent, supporting the ability of the model to describe the influence of the solvent and composition on lysozyme unfolding. Model predictions agreed fairly well with published differential scanning calorimetric data. The influence of hydroxylic additives on protein's conformational stability does not extend to any special property of these components, but to their ability to perturb the surface tension of water. This model can be used to interpret and correlate thermal unfolding data and to solve the problem of protein stabilization

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Estimation of the stability of globular proteins

Cited by 44 publications

References 26 publications

Isoenthalpic and isoentropic temperatures and the thermodynamics of protein denaturation.

Isoenthalpic and isoentropic temperatures and the thermodynamics of protein denaturation.

Destabilizing loop swaps in the CDRs of an immunoglobulin V_L domain

Molecular thermodynamics of heat‐induced protein unfolding in aqueous media

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Estimation of the stability of globular proteins

Cited by 44 publications

References 26 publications

Isoenthalpic and isoentropic temperatures and the thermodynamics of protein denaturation.

Isoenthalpic and isoentropic temperatures and the thermodynamics of protein denaturation.

Destabilizing loop swaps in the CDRs of an immunoglobulin VL domain

Molecular thermodynamics of heat‐induced protein unfolding in aqueous media

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Destabilizing loop swaps in the CDRs of an immunoglobulin V_L domain