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

Journal Article (Journal Article)

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 beta-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.

Full Text

Duke Authors

Cited Authors

  • Naus, S; Reipschläger, S; Wildeboer, D; Lichtenthaler, SF; Mitterreiter, S; Guan, Z; Moss, ML; Bartsch, JW

Published Date

  • March 2006

Published In

Volume / Issue

  • 387 / 3

Start / End Page

  • 337 - 346

PubMed ID

  • 16542157

International Standard Serial Number (ISSN)

  • 1431-6730

Digital Object Identifier (DOI)

  • 10.1515/BC.2006.045


  • eng

Conference Location

  • Germany