An infrared spectroscopic study of the interactions of carbohydrates with dried proteins.
Fourier-transform infrared spectroscopy was used to characterize the interaction of stabilizing carbohydrates with dried proteins. Freeze-drying of trehalose, lactose, and myo-inositol with lysozyme resulted in substantial alterations of the infrared spectra of the dried carbohydrates. In the fingerprint region (900-1500 cm-1), there were large shifts in the frequencies of bands, a decrease in absorbance, and a loss of band splitting. These effects mimic those of water on hydrated trehalose. Bands assigned to hydroxyl stretching modes (around 3350 cm-1) were decreased in intensity and shifted to higher frequencies in the presence of the protein. In complementary experiments, it was found that dehydration-induced shifts in the positions of amide I and amide II bands for lysozyme could be partially and fully reversed, respectively, when the protein was freeze-dried in the presence of either trehalose or lactose. In addition, the carboxylate band, which was not detectable in the protein dried without the sugar, was apparent when these sugars were present. myo-Inositol was less effective at shifting the amide bands, and the carboxylate band was not detected in the presence of this carbohydrate. Also tested was the concentration dependency of the carbohydrates' influence on the position of the amide II band for dried lysozyme. The results showed that the ability of a given concentration of a carbohydrate to shift this band back toward the position noted with the hydrated protein coincided, at least in the extreme cases, with the capacity of that same level of carbohydrate to preserve the activity of rabbit skeletal muscle phosphofructokinase during freeze-drying.(ABSTRACT TRUNCATED AT 250 WORDS)
Carpenter
J F
JF
Department of Zoology, University of California, Davis 95616.
Crowe
J H
JH
D002241:Carbohydrates; D004355:Drug Stability; D005612:Freeze Drying; D007294:Inositol; D007785:Lactose; D009113:Muramidase; D010732:Phosphofructokinase-1 / Q000378:metabolism; D012710:Serum Albumin, Bovine; D013055:Spectrophotometry, Infrared / Q000379:methods; D014199:Trehalose
D002241:Carbohydrates; D012710:Serum Albumin, Bovine; D007294:Inositol; D014199:Trehalose; D010732:Phosphofructokinase-1; D009113:Muramidase; D007785:Lactose