Hydrolysis of membrane-bound liver alkaline phosphatase by GPI-PLD requires bile salts.
Circulating liver plasma membrane fragments (LPMF) were purified from human serum by means of a monoclonal antileucine aminopeptidase antibody, AD-1. This was done by immunoaffinity chromatography or by incubating the sera with AD-1-coated nitrocellulose disks. Alkaline phosphatase (ALP, EC 3.1.3.1) is bound to these LPMF through a glycosylphosphatidylinositol (GPI) anchor and is referred to as membrane-bound liver ALP (Mem-LiALP). Low concentrations of Triton X-100 or high bile salt concentrations released GPI anchor-bearing LiALP (Anch-LiALP) from purified LPMF; once released, Anch-LiALP was slowly and progressively converted to hydrophilic dimeric LiALP [soluble LiALP (Sol-LiALP)], free from its GPI anchor. Low levels of GPI-specific phospholipase D (GPI-PLD) activity were measured in the pure LPMF. Apparently, this membrane-associated GPI-PLD was released by the action of detergents and contributed to the spontaneous conversion of Anch-LiALP to Sol-LiALP. In the absence of detergents, GPI-PLD had little effect on Mem-LiALP, both in purified form as well as in serum. In vitro, isolated Anch-LiALP was converted to Sol-LiALP by both GPI-specific phospholipase C and GPI-PLD. Sol-LiALP in serum, however, appeared to be the product of GPI-PLD activity only. Five- to tenfold higher concentrations of Triton X-100 were needed to release Anch-LiALP from LPMF in serum, compared with those required in a solution of purified LPMF. In serum, as well as in purified conditions, only a small range of detergent of bile salt concentrations permitted the conversion of Mem-LiALP to Sol-LiALP. A model is proposed for the release in the circulation of Mem-LiALP, Anch-LiALP, and Sol-LiALP, involving both LPMF-associated GPI-PLD and liver sinusoid bile salts.
Department of Nephrology-Hypertension, University of Antwerp, Belgium.
J T
JT
Deng
M F
MF
Hoylaerts
M E
ME
De Broe
V O
VO
van Hoof
D000469:Alkaline Phosphatase / Q000097:blood / Q000378:metabolism*; D001647:Bile Acids and Salts / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology; D002779:Cholestasis / Q000201:enzymology; D006801:Humans; D008099:Liver / Q000201:enzymology*; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D010739:Phospholipase D / Q000378:metabolism*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D012995:Solubility
D001647:Bile Acids and Salts; D000469:Alkaline Phosphatase; D010727:Phosphoric Diester Hydrolases; D010739:Phospholipase D; C065017:glycoprotein phospholipase D; D043265:Phosphatidylinositol Diacylglycerol-Lyase