Stabilisation and partial purification of Triton X-100 solubilised trihydroxycoprostanoyl-CoA synthetase from rat liver.
The stability of rat hepatic trihydroxycoprostanoyl-CoA syntethase was studied in its native membrane environment and after solubilisation by Triton X-100, and compared to that of choloyl-CoA synthetase. The lability of both delipidated enzymes could be suppressed by high concentrations of polyols such as sucrose and glucose. Addition of phospholipids to the assay mixtures was necessary to restore the activity of the stabilized enzymes. For further chromatographic separations, the addition of the hydrotrope Triton H-66 to the glucose-stabilized Triton X-100 solubilised synthetases improved their recovery on different matrices. Gel filtration revealed a native molecular mass of the Triton X-100/Triton H-66/protein micelles of 212 and 207 kDa for choloyl-CoA synthetase and trihydroxycoprostanoyl-CoA synthetase respectively.
Katholieke Universiteit Leuven, Department of Molecular Cell Biology, Belgium.
P P
PP
Van Veldhoven
S
S
Asselberghs
H J
HJ
Eyssen
G P
GP
Mannaerts
D000818:Animals; D002462:Cell Membrane / Q000737:chemistry / Q000201:enzymology; D002845:Chromatography / Q000379:methods; D002850:Chromatography, Gel; D003066:Coenzyme A Ligases / Q000737:chemistry* / Q000378:metabolism; D003902:Detergents / Q000737:chemistry; D004795:Enzyme Stability; D005947:Glucose / Q000737:chemistry; D008099:Liver / Q000737:chemistry / Q000201:enzymology*; D008297:Male; D017830:Octoxynol / Q000737:chemistry*; D010743:Phospholipids / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012995:Solubility
D003902:Detergents; D010743:Phospholipids; D017830:Octoxynol; D003066:Coenzyme A Ligases; C058701:trihydroxycoprostanoyl coenzyme A synthetase; C019890:choloyl-CoA synthetase; D005947:Glucose