Cleavage of the cell-surface O-sialoglycoproteins CD34, CD43, CD44, and CD45 by a novel glycoprotease from Pasteurella haemolytica.

The study of structural/functional characteristics of the cell-surface glycoproteins of leukocytes has led to a better understanding of the differentiation and maturation of hematopoietic cells. We have assessed the ability of a unique metalloprotease that is secreted by the bovine fibrinous pneumonia pathogen Pasteurella haemolytica, to cleave cell-surface glycoproteins expressed on human leukocytes. Biochemical analysis shows that the O-glycosylated cell surface Ag CD34, CD43 (leukosialin), CD44 (hyaluronic acid receptor), and CD45 (leukocyte common Ag), are all cleaved by this protease. Although these enzyme-sensitive structures contain N-linked glycans, they are all extensively glycosylated with O-linked carbohydrates, which are especially abundant on CD34 and CD43. In contrast, the glycoproteins CD18/11a,b,c (leukocyte integrins), CD71 (transferrin receptor), HLA class I, and 8A3 Ag, which contain N-linked glycans but no O-sialo-glycans, were resistant to the action of the enzyme. Inasmuch as previous studies using glycophorin A had indicated that the substrate specificity of this enzyme may be uniquely restricted to the cleavage of O-sialoglycoproteins, we have designated this activity, P. haemolytica glycoprotease. Immunofluorescence analysis with a variety of antibodies to different epitopes of the P. haemolytica glycoprotease-sensitive structures indicate that this enzyme may have widespread applications in epitope-mapping studies, and represents a novel tool with which to study structure/function relationships for O-sialoglycosylated cell-surface proteins. However, most significantly these results suggest that the P. haemolytica glycoprotease may be of use in the affinity purification and recovery of clinically important leukocyte subsets, such as primitive hematopoietic progenitors that express CD34.