Analysis of hydration water around human serum albumin using near-infrared spectroscopy

Abstract

Hydration plays a fundamental role in maintaining the structure and function of proteins. To get a generalized picture of hydrogen bond network of water surrounding human serum albumin (HSA), near-infrared (NIR) spectroscopy was adopted to explore the hydration induced structural changes of water with HSA concentration from 0.015 to 0.746 mmol/L. As HSA concentration increases, there was a nonlinear change in molar extinction coefficients inconsistent with Beer-Lambert law indicating the changes of hydration water induced by HSA and subsequently confirmed by the hydration number. The decreasing of hydration number with HSA concentration was explained by an overlapping hydration layer model. Resolution of the difference spectra with McCabe-Fisher method and aquaphotomics clearly differentiated the hydrogen bonding of hydration water around HSA. A comparison of resolved hydration spectra highlights that free hydrogen bonded water is present in the hydration layer. As the concentration increased, a more ordered hydrogen bonded water network forms around HSA. These measurements provide unique insight into the relationship between the hydration water and HSA, which is important for understanding the dynamics of protein solution in many biochemical processes, and may serve as a basis for the purification in production.