Quantitation of pH-induced Aggregation in Binary Protein Mixtures by Dielectric Spectroscopy
- 265 Downloads
This paper presents a quantitative approach for measuring pH-controlled protein aggregation using dielectric spectroscopy. The technique is demonstrated through two aggregation experiments, the first between β-lactoglobulin (β-Lg) and hen lysozyme (HENL) and the second between bovine serum albumin (BSA) and HENL. In both experiments, the formation of aggregates is strongly dependent on the solution pH and is clearly indicated by a decrease in the measured permittivity when the second protein is added. A quantifiable lower-bound on the ratio of proteins involved in the aggregation process is obtained from the permittivity spectra. Lower-bound aggregation ratios of 83 % for β-Lg/HENL at pH 6.0 and 48 % for BSA/HENL at pH 9.2 were consistent with turbidity measurements made on the same solutions.
KeywordsDielectric spectroscopy Protein aggregation pH
Bovine serum albumin
Protein data bank
We thank Shiul Khadka and Bryson Lanterman for initial construction of measurement cells and preliminary experiments. BLM was supported by a MIT Lincoln Laboratory Fellowship Award. This work was supported by funds from the BYU College of Engineering and Technology and the Office of Research & Creative Activities.
- 3.Choi I, Huh YS, Erickson D (2012) Microfluid Nanofluid (12):663–669Google Scholar
- 4.Cole KS, Cole RH (1941) J Chem Phys (9):341–351Google Scholar
- 5.Debye P (1929) Polar molecules. Chemical Catalog Company Inc, New YorkGoogle Scholar
- 11.Grant EH, Sheppard RJ, South GP (1978) Dielectric behaviour of biological molecules in solution. Oxford University Press, OxfordGoogle Scholar
- 19.MacKenzie H (1971) Milk proteins: chemistry and molecular biology, vol 2. Academic Press, New YorkGoogle Scholar
- 20.Mahler H, Jiskoot W (2011) Analysis of aggregates and particles in protein pharmaceuticals. Wiley, Hoboken NJGoogle Scholar
- 25.Mellor B, Kellis N, Mazzeo B (2011) In: 2011 annual international conference of the IEEE engineering in medicine and biology society, EMBC, Boston, MA, pp 3660–3663Google Scholar
- 35.Takashima S (1964) Biochim Biophys Acta Biophys Subj 79:531–538Google Scholar