A SDD1-like subtilase is exuded by tobacco roots

Hydroponically grown tobacco (Nicotiana tabacum L. cv. Samsun) roots exude proteases under non-stressed conditions. Ten different proteases could be distinguished by 2D-zymography of root exudate. The majority of the gelatinolytic activity was susceptible to serine protease inhibitors. One of the proteases could be assigned to an EST (SGN-P361478) by mass spectrometry of immune-purified root exudate. The sequence was completed by RACE-PCR and shows typical serine protease features of subtilase family S8A. Thermostability and SDS-insensitivity indicate a kinetically stable enzyme. Phylogenetic classification of this highly gelatinolytic subtilase showed SDD1 to be the closest relative in Arabidopsis thaliana (L. Heynh.). Even closer related protein sequences could be found in other distant plant genera indicating a high conservation of the subtilase. A 5-methyltetrahydropteroyltriglutamate-homocysteine methyltransferase-like protein and suberisation-associated anionic peroxidase-like protein were co-immune-purified and identified by mass spectrometry and may constitute potential interaction partners.