Temperature scanning FTIR analysis of secondary structures of proteins embedded in amorphous sugar matrix

J Pharm Sci. 2009 Sep;98(9):3088-98. doi: 10.1002/jps.21568.

Abstract

Heat-induced changes in secondary structures of five proteins (bovine serum albumin, BSA; human serum albumin, HSA; myoglobin; ribonuclease A, RNase A; and, beta-lactoglobulin, beta-Lg) in an amorphous sugar matrix were analyzed by temperature-scanning Fourier transform infrared spectroscopy to elucidate the mechanism of heat-induced conformational change of solid-phase proteins. Three sugars, trehalose, maltose, and dextran (MW 6000), were used. Loss of alpha-helices due to increasing temperature was observed for BSA, HSA, and myoglobin, which are rich in alpha-helices. RNase A showed a marked decrease in predominant secondary structural components (beta-sheet) with increasing temperature. However, no noticeable changes in the content of secondary structures, except for a slight loss of alpha-helices, were observed for beta-Lg, which is also beta-sheet-rich. These heat-induced conformational changes were significant at temperatures above the glass transition temperature. The heat-induced conformational change in BSA dried with sugar appeared time-independent and was clearly different from that due to dehydration and from the thermal conformational change for a solution of BSA. In particular, differences in secondary structural components that increased due to loss of alpha-helices were noted.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cattle
  • Dextrans / chemistry
  • Glucans / chemistry*
  • Humans
  • Hydrogen Bonding
  • Maltose / chemistry
  • Phase Transition
  • Protein Denaturation
  • Protein Structure, Secondary
  • Proteins / analysis*
  • Proteins / chemistry
  • Spectroscopy, Fourier Transform Infrared*
  • Temperature
  • Trehalose / chemistry

Substances

  • Dextrans
  • Glucans
  • Proteins
  • Maltose
  • Trehalose