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Two-Dimensional Near-Infrared Spectroscopy Study of Human Serum Albumin in Aqueous Solutions: Using Overtones and Combination Modes to Monitor Temperature-Dependent Changes in the Secondary Structure

2000; American Chemical Society; Volume: 104; Issue: 24 Linguagem: Inglês

10.1021/jp000537z

1520-6106

Yuqing Wu, Bogusława Czarnik‐Matusewicz, Koichi Murayama, Yukihiro Ozaki,

Spectroscopy and Quantum Chemical Studies

FT-NIR spectra were measured for human serum albumin (HSA) in aqueous solutions with concentrations of 1.0, 2.0, 3.0, 4.0, and 5.0 wt % over a temperature range of 45−80 °C. Concentration-perturbed two-dimensional (2D) correlation spectra were calculated for the spectra in the 7500−5500 cm-1 and 4900−4200 cm-1 regions at different temperatures. To investigate temperature-induced changes in the secondary structure and hydration, power spectra and slice spectra were calculated from the synchronous and asynchronous spectra, respectively. In the power spectra, a band near 4600 cm-1 due to the combination mode of amide B and amide II (amide B/II) shows an abrupt shift by 5 cm-1 between 58 and 60 °C, indicating that the secondary structure of HSA changes suddenly near 60 °C. Both the power and slice spectra in the 7500−5500 cm-1 region provide explicit evidence that the hydration changes markedly near 60 °C. A comparison of the temperature-dependent frequency shifts between the band near 4600 cm-1 due to amide B/II and that near 7000 cm-1 due to the combination mode of water reveals that the protein unfolding occurs almost in parallel with the change in the hydration. The present study demonstrates that the overtone and combination modes are very useful in monitoring subtle changes in protein dynamics.