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New insight into the pH-dependent conformational changes in bovine beta-lactoglobulin from Raman optical activity.

ABSTRACT: We have studied the conformation of beta-lactoglobulin in aqueous solution at room temperature over the pH range approximately 2.0-9.0 using vibrational Raman optical activity (ROA). The ROA spectra clearly show that the basic up and down beta-barrel core is preserved over the entire pH range, in agreement with other studies. However, from the shift of a sharp positive ROA band at approximately 1268 to approximately 1294 cm(-1) on going from pH values below that of the Tanford transition, which is centered at pH approximately 7.5, to values above, the Tanford transition appears to be associated with changes in the local conformations of residues in loop sequences possibly corresponding to a migration into the alpha-helical region of the Ramachandran surface from a nearby region. These changes may be related to those detected in X-ray crystal structures which revealed that the Tanford transition is associated with conformational changes in loops which form a doorway to the interior of the protein. The results illustrate how the ability of ROA to detect loop and turn structure separately from secondary structure is useful for studying conformational plasticity in proteins.

SUBMITTER: Blanch EW

PROVIDER: S-EPMC2144349 | biostudies-other | 1999 Jun

REPOSITORIES: biostudies-other

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New insight into the pH-dependent conformational changes in bovine beta-lactoglobulin from Raman optical activity.

Blanch E W EW Hecht L L Barron L D LD

Protein science : a publication of the Protein Society 19990601 6

We have studied the conformation of beta-lactoglobulin in aqueous solution at room temperature over the pH range approximately 2.0-9.0 using vibrational Raman optical activity (ROA). The ROA spectra clearly show that the basic up and down beta-barrel core is preserved over the entire pH range, in agreement with other studies. However, from the shift of a sharp positive ROA band at approximately 1268 to approximately 1294 cm(-1) on going from pH values below that of the Tanford transition, which ...[more]

PMID: 10386887

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Project description:Although bovine beta-lactoglobulin assumes a monomeric native structure at pH 3 in the absence of salt, the addition of salts stabilizes the dimer. Thermodynamics of the monomer-dimer equilibrium dependent on the salt concentration were studied by sedimentation equilibrium. The addition of NaCl, KCl, or guanidine hydrochloride below 1 M stabilized the dimer in a similar manner. On the other hand, NaClO(4) was more effective than other salts by about 20-fold, suggesting that anion binding is responsible for the salt-induced dimer formation, as observed for acid-unfolded proteins. The addition of guanidine hydrochloride at 5 M dissociated the dimer into monomers because of the denaturation of protein structure. In the presence of either NaCl or NaClO(4), the dimerization constant decreased with an increase in temperature, indicating that the enthalpy change (DeltaH(D)) of dimer formation is negative. The heat effect of the dimer formation was directly measured with an isothermal titration calorimeter by titrating the monomeric beta-lactoglobulin at pH 3.0 with NaClO(4). The net heat effects after subtraction of the heat of salt dilution, corresponding to DeltaH(D), were negative, and were consistent with those obtained by the sedimentation equilibrium. From the dependence of dimerization constant on temperature measured by sedimentation equilibrium, we estimated the DeltaH(D) value at 20 degrees C and the heat capacity change (DeltaC(p)) of dimer formation. In both NaCl and NaClO(4), the obtained DeltaC(p) value was negative, indicating the dominant role of burial of the hydrophobic surfaces upon dimer formation. The observed DeltaC(p) values were consistent with the calculated value from the X-ray dimeric structure using a method of accessible surface area. These results indicated that monomer-dimer equilibrium of beta-lactoglobulin at pH 3 is determined by a subtle balance of hydrophobic and electrostatic effects, which are modulated by the addition of salts or by changes in temperature.

Dataset Information

New insight into the pH-dependent conformational changes in bovine beta-lactoglobulin from Raman optical activity.

Publications

New insight into the pH-dependent conformational changes in bovine beta-lactoglobulin from Raman optical activity.

Similar Datasets

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