Conformational changes and inactivation of calf intestinal alkaline phosphatase in trifluoroethanol solutions.
The changes in activity and unfolding of calf intestinal alkaline phosphatase (CIP) during denaturation in different concentrations of trifluoroethanol (TFE) have been investigated by far-ultraviolet circular dichroism and fluorescence emission spectra. Unfolding and activation rate constants were measured and compared, the activation and inactivation courses were much faster than that of unfolding, which suggests that the active site of CIP containing two zinc ions and one magnesium ion is situated in a limited and flexible region of the enzyme molecule that is more fragile to the denaturant than the protein as a whole. However, compared to other metalloenzymes, CIP is inactivated at higher concentrations of TFE as denaturant.
Zhang
Y X
YX
Department of Chemistry, Capital University of Medical Sciences, Beijing, People's Republic of China.
Zhu
Y
Y
Zhou
H M
HM
D000469:Alkaline Phosphatase / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism; D000581:Amidohydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002942:Circular Dichroism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D007422:Intestines / Q000201:enzymology*; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism; D008667:Metalloproteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D017433:Protein Structure, Secondary / Q000187:drug effects; D013050:Spectrometry, Fluorescence; D014270:Trifluoroethanol / Q000494:pharmacology*; D014466:Ultraviolet Rays; D015032:Zinc / Q000378:metabolism
D008667:Metalloproteins; D014270:Trifluoroethanol; D000469:Alkaline Phosphatase; D000581:Amidohydrolases; C019672:aminoacylase I; D008274:Magnesium; D015032:Zinc