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Identification of novel dipeptidyl peptidase 9 substrates by two‐dimensional differential in‐gel electrophoresis
Author(s) -
Zhang Hui,
Maqsudi Sadiqa,
Rainczuk Adam,
Duffield Nadine,
Lawrence Josie,
Keane Fiona M.,
JustaSchuch Daniela,
GeissFriedlander Ruth,
Gorrell Mark D.,
Stephens Andrew N.
Publication year - 2015
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.13371
Subject(s) - dipeptidyl peptidase , dipeptidyl peptidase 4 , serine protease , gel electrophoresis , chemistry , serine , microbiology and biotechnology , biochemistry , cleavage (geology) , biology , enzyme , protease , type 2 diabetes , diabetes mellitus , endocrinology , paleontology , fracture (geology)
Dipeptidyl peptidase 9 ( DPP 9) is a member of the S9B/ DPPIV ( DPP 4) serine protease family, which cleaves N‐terminal dipeptides at an Xaa‐Pro consensus motif. Cytoplasmic DPP 9 has roles in epidermal growth factor signalling and in antigen processing, whilst the role of the recently discovered nuclear form of DPP 9 is unknown. Mice lacking DPP 9 proteolytic activity die as neonates. We applied a modified 2D differential in‐gel electrophoresis approach to identify novel DPP 9 substrates, using mouse embryonic fibroblasts lacking endogenous DPP 9 activity. A total of 111 potential new DPP 9 substrates were identified, with nine proteins/peptides confirmed as DPP 9 substrates by MALDI ‐ TOF or immunoblotting. Moreover, we also identified the dipeptide Val‐Ala as a consensus site for DPP 9 cleavage that was not recognized by DPP 8, suggesting different in vivo roles for these closely related enzymes. The relative kinetics for the cleavage of these nine candidate substrates by DPP 9, DPP 8 and DPP 4 were determined. This is the first identification of DPP 9 substrates from cells lacking endogenous DPP 9 activity. These data greatly expand the potential roles of DPP 9 and suggest different in vivo roles for DPP 9 and DPP 8.