Purification and characterization of human bone tartrate-resistant acid phosphatase.
Tartrate-resistant acid phosphatase (TRAP) is a histochemical marker for osteoclasts, the multinucleated bone resorbing cell. This type 5 acid phosphatase has been purified 500-fold from human bone by three chromatographic steps: cation exchange, gel filtration, and HPLC cation exchange. Like most other TRAPs isolated, it is a basic glycoprotein of a molecular weight about 33,000. Its pH optimum Km, and Vmax for p-nitrophenyl phosphate are 5.7, 0.8 mM, and 12 units/mg, respectively. Human bone TRAP hydrolyzes aryl phosphates, nucleoside di- and triphosphates, pyrophosphate, and phosphoproteins. It is activated by mild reducing agents but inhibited by molybdate, fluoride, arsenate, phosphate, and dithionite. Its activity is not inhibited by tartrate, a feature that distinguishes it from other acid phosphatases. Sodium etridonate, the bisphosphonate used clinically to reduce bone resorption, is a relatively poor inhibitor of bone TRAP. Human bone TRAP is immunologically related to the porcine uterine secretory TRAP, uteroferrin. Monospecific rabbit antibodies to the bone TRAP have been immunopurified by using affinity chromatography with uteroferrin immobilized on Sepharose and can be used to detect low amounts of the enzyme in a simple dot-blot assay.
Allen
S H
SH
Department of Medicine, University of Missouri, Columbia 65212.
Nuttleman
P R
PR
Ketcham
C M
CM
Roberts
R M
RM
D000135:Acid Phosphatase / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D015415:Biomarkers / Q000032:analysis; D001842:Bone and Bones / Q000201:enzymology*; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007120:Immunochemistry; D008970:Molecular Weight; D013379:Substrate Specificity; D013644:Tartrates / Q000494:pharmacology
D015415:Biomarkers; D013644:Tartrates; D000135:Acid Phosphatase; C029768:tartaric acid