Purification and properties of the native form of the purple acid phosphatase from bovine spleen.
The purple acid phosphatase (PAP) from bovine spleen has been shown to exist as a single ca. 36-kDa polypeptide in intact spleen tissue. The previously isolated microheterogeneous complex of 15-kDa and 23- or 21-kDa subunits appears to arise from proteolytic cleavage of an exposed, highly variable loop in the polypeptide chain. Small amounts of a single polypeptide form, presumed to be the native form of the enzyme, have been obtained; this has permitted its optical and EPR spectra and fundamental kinetic properties to be determined. The most notable difference between the native and two-subunit forms of PAP is a ca. 3-fold higher enzymatic activity for the latter, which is due to a simple increase in Vmax. The two forms are very similar spectroscopically and chemically and appear to differ only in the loss of a highly antigenic ca. five amino acid segment of the polypeptide between positions 155 and 160 but not in NH2-terminal sequence or in carbohydrate content. Analysis of published sequence data suggests that the existence of an exposed highly antigenic loop at positions corresponding to 155-161 of the spleen PAP sequence is a relatively general feature of PAP's. Trypsin and chymotrypsin cleave both bovine spleen PAP and uteroferrin, apparently in this region, with significant enhancement of enzymatic activity.
Department of Chemistry, University of Virginia, Charlottesville 22901.
J L
JL
Orlando
T
T
Zirino
B J
BJ
Quirk
B A
BA
Averill
D000135:Acid Phosphatase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D015153:Blotting, Western; D002241:Carbohydrates / Q000032:analysis; D002417:Cattle; D002942:Circular Dichroism; D004578:Electron Spin Resonance Spectroscopy; D010450:Endopeptidases / Q000494:pharmacology; D007527:Isoenzymes; D007700:Kinetics; D008667:Metalloproteins / Q000378:metabolism; D008969:Molecular Sequence Data; D008982:Molybdenum / Q000494:pharmacology; D010710:Phosphates / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D017421:Sequence Analysis; D013053:Spectrophotometry; D013154:Spleen / Q000201:enzymology*; D000071681:Tartrate-Resistant Acid Phosphatase
D000596:Amino Acids; D002241:Carbohydrates; D007527:Isoenzymes; D008667:Metalloproteins; D010710:Phosphates; D011480:Protease Inhibitors; C044659:molybdate; D008982:Molybdenum; D000135:Acid Phosphatase; D000071681:Tartrate-Resistant Acid Phosphatase; D010450:Endopeptidases