The circulatory protein, human serum albumin (HSA), is widely used as a model protein for the study of protein structure. In this work, the structures of human serum albumin in aqueous solutions are studied using temperature-dependent near-infrared (NIR) spectroscopy with the aid of continuous wavelet transform (CWT). Near-infrared spectra of human serum albumin solutions with different concentrations were measured over a temperature range of 30–85 ℃. Then, continuous wavelet transform was performed on the spectra to enhance the resolution. As a result of the resolution enhancement, spectral bands around 4361, 4521, 4600 and 4260 cm−1 were extracted from the overlapping low-resolution signals. The four bands can be assigned to the protein structures of α-helix, β-sheet, an intermediate state and side chains, respectively. The variations in intensity of the bands around 4361 and 4521 cm−1 with temperature show that the increase of temperature leads to the loss of α-helical structure but the formation of β-sheet, and the denaturation temperature of human serum albumin is about 55 ℃. The variation of the band around 4600 cm−1 indicates that the temperature-induced unfolding process of human serum albumin occurs through a stable intermediate state, and a significant change in the microenvironment of the side chains about 63 ℃ is observed from the variation of the band around 4260 cm−1. On the other hand, the transformed spectra in the region of 8000–5600 cm−1 provide an explicit evidence for the structural changes of water during the process of protein denaturation, and the unfolding process of HSA can be reflected by these changes.
Selective detection of human serum albumin by near infrared emissive fluorophores: Insights into structure-property relationship
