Liver-like alkaline phosphatase in the tissue-unspecific type enzyme found in rabbit organs.

Rabbit liver and kidney tissues are known to produce an intestinal-like alkaline phosphatase (IAP-like enzyme) as a dominant isozyme, with a minor isozyme of tissue-unspecific type (UAP), unlike humans and other mammalians. We investigated immunohistochemically and biochemically these unique isozymes in the rabbit liver and bone, and compared them with the human isozyme. In rabbit liver, UAP was found to be localized only in the apical part of the membrane of cells lining the bile duct, whereas IAP-like enzyme was found in the sinusoidal membrane of hepatocytes. Rabbit liver UAP was separated from IAP-like enzyme by DEAE-cellulose column chromatography. Rabbit bone tissue contained only one UAP isozyme. The two UAPs were biochemically and physicochemically compared with human liver AP. Both UAPs reacted with an anti-human liver AP monoclonal antibody, not with an anti-human bone AP monoclonal antibody, indicating that both enzymes have the same antigenicity as human liver AP. Rabbit liver and bone UAPs had similar N-linked sugar-chain heterogeneities to the respective human enzymes. In addition, rabbit bone AP also had an O-linked sugar chain, as did human bone AP, unlike rabbit and human liver APs.

Intestinal alkaline phosphatase isoforms in rabbit tissues differ in glycosylation patterns.

OBJECTIVES: In patients with advanced liver cirrhosis or chronic nephritis, an intestinal alkaline phosphatase (IAP)-like enzyme is ectopically expressed in the liver or kidney. In this study, we used rabbit organs as a human pathological model, because the rabbit liver or kidney expresses an IAP-like enzyme as the predominant isozyme, unlike humans. METHODS: IAP and the IAP-like enzyme were purified from rabbit intestine and kidney, respectively, by immunoaffinity chromatography using a monoclonal antihuman IAP antibody. Some properties of the IAP and IAP-like enzyme expressed in rabbit organs are compared. RESULTS: Some of their catalytic and physicochemical properties differed. In particular, the net charge, molecular mass, and hydrophobicity of IAP from rabbit intestine was slightly different from the IAP-like enzyme from rabbit kidney. There was a difference in the sugar chain structure between the two enzymes according to the results of lectin affinity chromatography, and part of the peptide maps differed slightly. However, there was no difference in the peptide maps after treatment with endo-N-acetylglucosaminidase F. CONCLUSIONS: The primary structures of IAP and the IAP-like enzyme are basically similar, except for the glycosylation process of the respective AP isozymes.

Intestinal type alkaline phosphatase hyperphosphatasemia associated with liver cirrhosis.

Hyperphosphatasemia due to increased intestinal type serum alkaline phosphatase was noted in a 48-year-old male who had asymptomatic liver cirrhosis. The alkaline phosphatase activity in the serum was 828 U/l (our reference range in adults: 57-194 U/l), 94% of which was of the intestinal type as measured by an immunoprecipitation method. The intestinal component of alkaline phosphatase was separated into two major and some minor components using electrophoresis and isoelectrofocusing. One of the major components had similar mobility to that of a standard intestinal enzyme purified from adult intestine. The components were heat-labile and neuraminidase-resistant. Serial lectin affinity chromatography, however, indicated that sugar chain compositions of the alkaline phosphatase were different from those of the standard tissue intestinal enzyme. These results and further enzymological studies suggest that the patient's serum alkaline phosphatase basically consisted of several intestine-like isoforms.

Purification and partial characterization of intestinal-like alkaline phosphatase in rabbit kidney.

Two types of alkaline phosphatase (AP) isozymes in rabbit kidney, a major intestinal-like type and a minor tissue-unspecific type, have been identified. The former enzyme was purified from rabbit kidney by immunoaffinity chromatography using monoclonal anti-human intestinal AP antibody. The purified enzyme yielded a single protein band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the apparent molecular size of its monomer subunit was found to be 72,000. Three amino acid residues within the first 16 N-terminal amino acid residues were different in purified AP and human intestinal AP. Although the rabbit enzyme possessed some peptide bands identical to those of human adult intestinal AP after Staphylococcus aureus V8 protease digestion, the enzyme did not react with monoclonal antibody against human adult intestinal AP alone, whereas it did react with monoclonal antibody against both human adult and fetal intestinal APs. The affinity of the enzyme for concanavalin A was identical to that of the fetal intestinal AP, but different from that of the adult enzyme. These results indicate that the antigenicity and certain properties of purified rabbit AP are more like those of human fetal intestinal AP or Kasahara isozyme, so-called intestinal-like AP, than like human adult intestinal AP.