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Biological Chemistry

Editor-in-Chief: Brüne, Bernhard

Editorial Board Member: Buchner, Johannes / Ludwig, Stephan / Sies, Helmut / Turk, Boris / Wittinghofer, Alfred

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Identification of candidate substrates for ectodomain shedding by the metalloprotease-disintegrin ADAM8

Silvia Naus1 / Simone Reipschläger2 / Dirk Wildeboer3 / Stefan F. Lichtenthaler4 / Stefan Mitterreiter5 / Ziqiang Guan6 / Marcia L. Moss7 / Jörg W. Bartsch8









Corresponding author

Citation Information: Biological Chemistry. Volume 387, Issue 3, Pages 337–346, ISSN (Online) 1437-4315, ISSN (Print) 1431-6730, DOI: 10.1515/BC.2006.045, March 2006

Publication History

October 24, 2005
December 14, 2005
Published Online:


ADAM proteases are type I transmembrane proteins with extracellular metalloprotease domains. As for most ADAM family members, ADAM8 (CD156a, MS2) is involved in ectodomain shedding of membrane proteins and is linked to inflammation and neurodegeneration. To identify potential substrates released under these pathologic conditions, we screened 10-mer peptides representing amino acid sequences from extracellular domains of various membrane proteins using the ProteaseSpot™ system. A soluble ADAM8 protease containing a pro- and metalloprotease domain was expressed in E. coli and purified as active protease owing to autocatalytic prodomain removal. From 34 peptides tested in the peptide cleavage assay, significant cleavage by soluble ADAM8 was observed for 14 peptides representing membrane proteins with functions in inflammation and neurodegeneration, among them the β-amyloid precursor protein (APP). The in vivo relevance of the ProteaseSpot™ method was confirmed by cleavage of full-length APP with ADAM8 in human embryonic kidney 293 cells expressing tagged APP. ADAM8 cleaved APP with similar efficiency as ADAM10, whereas the inactive ADAM8 mutant did not. Exchanging amino acids at defined positions in the cleavage sequence of myelin basic protein (MBP) revealed sequence criteria for ADAM8 cleavage. Taken together, the results allowed us to identify novel candidate substrates that could be cleaved by ADAM8 in vivo under pathologic conditions.

Keywords: ADAM protease; candidate substrates; ectodomain shedding; fluorescence assay; peptide cleavage

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