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cDNA encoding a functional feline liver/bone/kidney-type alkaline phosphatase

A K Ghosh1, J I Mullins

  • 1Department of Microbiology and Immunology, Stanford University School of Medicine, California 94305-5402, USA.

Archives of Biochemistry and Biophysics
|September 10, 1995
PubMed
Summary

Feline alkaline phosphatase (FeALP) was identified as a functional enzyme, but it does not act as a receptor for feline leukemia virus subgroup-A (FeLV-A). This study characterized the FeALP gene and its protein product.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Virology

Background:

  • Feline alkaline phosphatase (FeALP) was initially copurified with a putative feline leukemia virus subgroup-A (FeLV-A) receptor.
  • The purified protein's sequence showed homology to mammalian liver/bone/kidney (L/B/K)-type alkaline phosphatases.

Purpose of the Study:

  • To clone and characterize the feline L/B/K-type alkaline phosphatase (FeALP) gene.
  • To determine if FeALP functions as the FeLV-A receptor.

Main Methods:

  • Polymerase chain reaction (PCR) amplification using degenerate oligonucleotides.
  • Screening of a feline T-lymphocyte cDNA library to isolate the FeALP cDNA clone.
  • Gene expression in murine and mink lung fibroblast cell lines.
  • Biochemical assays including enzymatic activity measurement, immunoblotting, and phospholipase-C treatment.

Related Experiment Videos

  • Northern blot analysis to detect mRNA species.

    • Main Results:

    • A 2127-bp cDNA encoding a 524-amino-acid protein homologous to mammalian L/B/K-type ALP was isolated.
    • Expressed FeALP exhibited enzymatic activity and was localized to the cell surface via a phosphatidylinositol-glycan (PIG) anchor.
    • FeALP did not facilitate FeLV-A attachment or entry into cells, indicating it is not the viral receptor.

    Conclusions:

    • The cloned cDNA represents a functional feline L/B/K-type alkaline phosphatase.
    • FeALP is expressed on the cell surface and anchored via a PIG linkage.
    • FeALP does not serve as the receptor for feline leukemia virus subgroup-A.