Investigation of bovine serum albumin denaturation using ultrasonic spectroscopy
journal contributionposted on 04.12.2012, 14:26 authored by Malcolm J.W. Povey, Jonathan D. Moore, Julian Braybrook, Howard Simons, Ron Belchamber, Meera Raganathan, Valerie PinfieldValerie Pinfield
The ability of ultrasound spectroscopy to characterise protein denaturation at relatively high concentrations and under conditions found in foods, is examined. Measurement of longitudinal sound velocity against concentration and frequency (20-160 MHz) for the bovine serum albumin monomer at pH 7.0 gave a frequency independent value for molecular compressibility of at 25 °C, corresponding to a sound velocity for the BSA molecule of 1920 ms-1. At 160 MHz, the longitudinal sound attenuation in BSA molecules is ~5200 Npm-1, a factor of 10 higher than in water. The excess attenuation of the solution over water was nearly 90 Npm-1 at the highest measured volume fraction of 0.03 (or 3% v/v). Concentration-dependent ultrasound velocity (20 - 160 MHz) and attenuation (2 - 120 MHz) spectra were obtained over time for heated bovine serum albumin (BSA) solutions up to 40 mg/mL at neutral pH and at 25 °C. An acoustic scattering model was used which considered the solute molecules as scatterers of ultrasound, to determine the molecules' sound velocity, compressibility, and attenuation properties. Mild heat treatment caused the molecule to organise into dimers and trimers, without change in sound velocity; implying that there is little or no change in secondary structure. Changes in attenuation spectra correlated with estimated molecular weight as determined through DLS and SEC measurements. During oligomerisation, the BSA molecules continue to behave acoustically as monomers. Under severe heat treatment, BSA rapidly suffered irreversible denaturation and gelation occurred which affected both ultrasound attenuation spectra and the velocity of sound, consistent with significant molecular conformation changes and/or molecule-molecule interactions.
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