Unusual thermal stabilities of some proteins and enzymes bound in the galleries of layered α-Zr(IV)phosphate/phosphonates

Abstract

Significant improvements in the thermal stabilities of proteins and enzymes (increase in denaturation temperatures as large as 33 °C) bound in the galleries of crystalline α-Zr(IV) phosphate (α-Zr(PO 4) 2·H 2O, abbreviated as α-ZrP) and α-Zr(IV) phosphonates (α-Zr(RPO 3) 2·H 2O, RCH 2COOH, CH 2CH 2COOH, abbreviated as ZrCMP, and ZrCEP, respectively) are demonstrated, in specific cases. The thermal stabilities are monitored using attenuated total reflection Fourier transform infrared spectroscopy (ATR FTIR), powder X-ray diffraction, absorption, and fluorescence methods. The amide I and amide II vibrational bands of the protein/inorganic materials are monitored in ATR FTIR studies, as a function of temperature, and these indicated sharp shifts in the band positions at specific temperatures. These shifts served as markers for protein denaturation. The denaturation temperatures of met hemoglobin/α-ZrP, glucose oxidase/α-ZrRP (ROH, CH 2CH 2COOH), and Cytochrome c/α-ZrRP (ROH, CH 2CH 2COOH), for example, are >95, 55, 65, 100, and 75 °C, respectively. These values are greater than the denaturation temperatures of the corresponding free proteins in the solid state (70, 50, and 67 °C, respectively). Proteins bound to α-ZrP, and α-ZrCEP, in general, indicated stabilities that are greater than or comparable to those of the free proteins, while binding to α-ZrCMP resulted in diminished stabilities except in case of Hb. The interactions of the surface functions of the solid matrix with the bound protein, therefore, are crucial in determining the bound protein stability. The improved stabilities observed in specific cases are welcome changes for applications of these materials in biocatalysis, biosensors, and biomedical devices.