J. Biol. Chem., Vol. 258, Issue 17, 10283-10288, Sep, 1983
The immunological and structural comparisons of deoxyribonucleases I. Glycosylation differences between bovine pancreatic and parotid deoxyribonucleases
A Abe and TH Liao
A rabbit antiserum against bovine pancreatic DNase A is used to study the
immunological reaction of DNases I. As shown by double immunodiffusion,
bovine pancreatic DNases A, B, C, and D are immunologically identical, so
are DNases from bovine pancreas and parotid and from ovine pancreas. These
DNases also behave similarly in immunotitration of DNase activity and all
are tightly bound to the immunoaffinity medium, requiring an acidic buffer
with 10% ammonium sulfate to dissociate. On the other hand, porcine
pancreatic and malted barley DNases that do not form precipitin lines
remain active in solution with the antibody; however, in spite of the lack
of inhibition these DNases are retarded (but not tightly bound) in
immunoaffinity chromatography, suggesting interaction with the antibody. In
thin layer isoelectric focusing, the parotid DNase, purified with the
immunoaffinity technique, shows only two major active components whose
isoelectric points correspond to those of DNases A and C of bovine
pancreas. As estimated by sodium dodecyl sulfate-polyacrylamide gel
electrophoresis, the molecular weight of parotid DNase is 34,000,
approximately 3,000 more than that of the pancreatic enzyme. However, both
parotid and pancreatic DNases have the same NH2-terminal leucine, an
identical COOH-terminal amino acid sequence, nearly identical amino acid
compositions, and almost the same peptide maps. The molecular weight
difference is due to differences in the carbohydrate side chains. Results
of peptide analyses indicate that parotid DNase contains two glycopeptides;
pancreatic DNase has only one. In addition, both parotid glycopeptides
contain glucosamine and galactosamine while the pancreatic glycopeptide has
only glucosamine.
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