Purification and characterization of amphiphilic trehalase from rabbit small intestine.
Rabbit intestinal trehalase (alpha,alpha-trehalose glucohydrolase, EC 3.2.1.28) was solubilized with Triton X-100 and purified in the presence of EDTA. The purified enzyme was homogeneous on polyacrylamide gel electrophoresis in the presence of Triton X-100 or SDS. It showed amphiphilic properties on gel filtration. polyacrylamide gel electrophoresis, charge-shift electrophoresis and phenyl-Sepharose chromatography. Its molecular weight was estimated to be about 330 000 by gel filtration under nondenaturing conditions and in the presence of Triton X-100, the value being in satisfactory agreement with the sum of the weight of one Triton X-100 micelle and twice the molecular weight (105 000) of purified hydrophilic trehalase which had been deprived of the anchor segment. The two purified trehalases gave almost the same molecular weights (about 75 000) on SDS-polyacrylamide gel electrophoresis. These results suggest that intestinal trehalase consists of two subunits with a molecular weight of 75 000 and that its anchor segment is small (less than 5000). Triton X-100 extracts freshly prepared from intestinal microvilli essentially showed one form of trehalase, which behaved on phenyl-Sepharose and Con A-Sepharose chromatography in the same manner as purified amphiphilic trehalase.
Yokota
K
K
Nishi
Y
Y
Takesue
Y
Y
D000818:Animals; D003902:Detergents; D007421:Intestine, Small / Q000201:enzymology*; D007527:Isoenzymes / Q000302:isolation & purification; D046911:Macromolecular Substances; D008823:Micelles; D008871:Microvilli / Q000201:enzymology*; D008970:Molecular Weight; D017830:Octoxynol; D011092:Polyethylene Glycols; D011817:Rabbits; D014198:Trehalase / Q000302:isolation & purification* / Q000378:metabolism
D003902:Detergents; D007527:Isoenzymes; D046911:Macromolecular Substances; D008823:Micelles; D011092:Polyethylene Glycols; D017830:Octoxynol; D014198:Trehalase