Characterization of a tartrate-resistant acid phosphatase (ATPase) from rat bone: hydrodynamic properties and N-terminal amino acid sequence.
Certain physicochemical properties of rat bone tartrate-resistant acid ATPase (TrATPase), including the size and shape of the enzyme, potential subunit composition, and detergent binding, have been elucidated. SDS-polyacrylamide gel electrophoresis in combination with immunoblot analysis showed that the bone TrATPase has a molecular weight of 33,000 D and is composed of disulfide-linked polypeptides of 20,000 and 16,000 D. The enzyme contains 1.7 mol Fe per mol enzyme. Hydrodynamic studies allowed calculation of the Stokes radius (24 A), the sedimentation coefficient (3.19S), the partial specific volume (0.748 ml/g), the frictional ratio (0.995), and the axial ratio (1.0). The amount of detergent bound to the protein was determined to 4 mol of Triton X-100 per mol enzyme. The molecular weight of bone TrATPase derived from these parameters was 31,900 D. N-terminal amino acid sequence analysis of the Mr 20,000 subunit indicated a high degree of similarity with TRAP enzymes from spleen, uterus, placenta, hairy cell leukemia, and osteoclastoma. It is concluded that rat bone TrATPase belongs to the type 5 (tartrate-resistant and purple) acid phosphatase family. The similarities in the N-terminal amino acid sequences, iron content, and physicochemical properties of TRAP enzymes indicate a close structural relationship between type 5 acid phosphatases expressed in different tissues. The findings that TrATPase has a spherical shape and binds low amounts of detergent suggest that the enzyme is a soluble protein, compatible with the view that TrATPase is secreted by the osteoclast.
Ek-Rylander
B
B
Department of Oral Pathology, Karolinska Institute, Huddinge, Sweden.
Bergman
T
T
Andersson
G
G
D000135:Acid Phosphatase / Q000032:analysis*; D000251:Adenosine Triphosphatases / Q000032:analysis*; D000595:Amino Acid Sequence; D000818:Animals; D001842:Bone and Bones / Q000201:enzymology*; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D015151:Immunoblotting; D007501:Iron / Q000032:analysis; D007527:Isoenzymes / Q000032:analysis*; D008969:Molecular Sequence Data; D051381:Rats; D012689:Sequence Homology, Nucleic Acid; D013644:Tartrates / Q000494:pharmacology*
D007527:Isoenzymes; D013644:Tartrates; D007501:Iron; D000135:Acid Phosphatase; D000251:Adenosine Triphosphatases