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- Title
The participation of phytocystatin TrcC-4 in the activity regulation of EP8, the main prolamin degrading cysteine endopeptidase in triticale seeds.
- Authors
Szewińska, Joanna; Prabucka, Beata; Krawczyk, Mariusz; Mielecki, Marcin; Bielawski, Wiesław
- Abstract
Phytocystatins (PCs) are protein inhibitors of endogenous plant endopeptidases and exogenous pathogen proteinases. We have previously described the protein inhibitor TrcC-4, which is probably involved in the control of protein degradation during triticale seeds germination. The occurrence of the LARFAVXEHN motif supports the TrcC-4 designation as a PC. In this paper TrcC- 4 was expressed in Escherichia coli using the pET28 expression vector. TrcC-4(6×His) was purified by affinity chromatography with a single step of purification. Western blot analysis showed the presence of TrcC-4 in both developing and germinating triticale seeds. TrcC-4 protein level was higher both in scutellum of germinating seeds and in developing grains of the triticale cultivar more resistant to pre-harvest sprouting (Zorro) than in a less resistant one (Disco). Furthermore it was demonstrated that the activity of EP8, cysteine endopeptidase responsible for the mass hydrolysis of prolamin during germination, is inhibited by TrcC-4(6×His), as confirmed by native PAGE with gliadin as a substrate. These results suggest that phytocystatin TrcC-4 controls the activity of cysteine endopeptidases involved in germination and, thus, is potentially involved in pre-harvest sprouting.
- Subjects
ENDOPEPTIDASES; CYSTEINE proteinases; PROTEOLYTIC enzymes; GERMINATION; ESCHERICHIA coli
- Publication
Plant Growth Regulation, 2013, Vol 69, Issue 2, p131
- ISSN
0167-6903
- Publication type
Article
- DOI
10.1007/s10725-012-9756-5