MONOCLONAL PROTEINASE 3 ANTIBODIES BINDING AT THE ACTIVE SITE

Activity: Talks and presentationsTalks and presentations in private or public companies

Description

M.Z. Hansen1, R. Bennike1, T. Ravnsborg2, C. Schou1, G. Houen1, P. Højrup2
1Department of Clinical Biochemistry and Immunology, Statens Serum Institut, Copenhagen, 2Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark

Proteinase 3 (PR3) is a neutrophil granulocyte enzyme involved in host antimicrobial defence.
We have characterized the glycosylation of PR3 and its association with alfa-defensins. Furthermore, we have characterised three monoclonal antibodies (mabs) against PR3 and their dependency on PR3 structure and modifications.
The three mabs (4A3, 4A5, 6A6) bound PR3 both in its native and denatured forms, provided the disulphide bridges were intact. Denatured PR3 with reduced disulphide bridges was not recognized by any of the three mabs. Alfa-1-antitrypsin (AT) bound purified human neutrophil granulocyte PR3 and inhibited its proteolytic activity towards a small synthetic peptide substrate and a large protein substrate (casein). AT also inhibited binding of the three mabs to PR3, indicating that they bind in a region affected by AT binding. Screening of overlapping synthetic peptides covering the whole PR3 sequence showed binding to peptides from the active site area, which was further investigated by a combination of chemical surface labelling and mass spectrometry. However, the mabs did not inhibit PR3 proteolytic activity, showing that they bind close to the active site without restricting access to the substrate cleft and the catalytic triad.

Period13. May 2012
Event title8th International Congress on Autoimmunity
Event typeConference
Conference number8
LocationGranada, Spain