Dual specificities of the glyoxysomal/peroxisomal processing protease Deg15 in higher plants. Hierl Georg Lück Carsten Helm Michael et al. 2007 Jul. Proc. Natl. Acad. Sci. U.S.A. 104(27):11501-6. Arnold Georg J Adamska Iwona Fröhlich Thomas 17592111 11501-6 Gietl Christine 104 Glyoxysomes are a subclass of peroxisomes involved in lipid mobilization. Two distinct peroxisomal targeting signals (PTSs), the C-terminal PTS1 and the N-terminal PTS2, are defined. Processing of the PTS2 on protein import is conserved in higher eukaryotes. The cleavage site typically contains a Cys at P1 or P2. We purified the glyoxysomal processing protease (GPP) from the fat-storing cotyledons of watermelon (Citrullus vulgaris) by column chromatography, preparative native isoelectric focusing, and 2D PAGE. The GPP appears in two forms, a 72-kDa monomer and a 144-kDa dimer, which are in equilibrium with one another. The equilibrium is shifted on Ca(2+) removal toward the monomer and on Ca(2+) addition toward the dimer. The monomer is a general degrading protease and is activated by denatured proteins. The dimer constitutes the processing protease because the substrate specificity proven for the monomer (Phi-Arg/Lys downward arrow) is different from the processing substrate specificity (Cys-Xxx downward arrow/Xxx-Cys downward arrow) found with the mixture of monomer and dimer. The Arabidopsis genome analysis disclosed three proteases predicted to be in peroxisomes, a Deg-protease, a pitrilysin-like metallopeptidase, and a Lon-protease. Specific antibodies against the peroxisomal Deg-protease from Arabidopsis (Deg15) identify the watermelon GPP as a Deg15. A knockout mutation in the DEG15 gene of Arabidopsis (At1g28320) prevents processing of the glyoxysomal malate dehydrogenase precursor to the mature form. Thus, the GPP/Deg15 belongs to a group of trypsin-like serine proteases with Escherichia coli DegP as a prototype. Nevertheless, the GPP/Deg15 possesses specific characteristics and is therefore a new subgroup within the Deg proteases. Prestele Jakob 27 Lottspeich Friedrich 2007 Jul Huesgen Pitter F Helm Michael Görg Angelika