The fungicidal type I chitinases contribute to the defense response of plants against pathogens. Two tobacco chitinases represent a different class of hydroxyproline-containing proteins. Hydroxyproline-rich proteins are predominantly extracellular, structural glycoproteins proteins that lack enzymatic activity and contain many hydroxyproline residues. In contrast, type I chitinases are vacuolar enzymes. They are not glycosylated and contain a small number of hydroxyproline residues restricted to a single, short peptide sequence.
Aspergillus fumigatus grows well at neutral and acidic pH in a medium containing protein as the sole nitrogen source by secreting two different sets of proteases. Neutral pH favors the secretion of neutral and alkaline endoproteases, leucine aminopeptidases (Laps) which are nonspecific monoaminopeptidases, and an X-prolyl dipeptidase (DppIV). Acidic pH environment promotes the secretion of an aspartic endoprotease of pepsin family (Pep1) and tripeptidyl-peptidases of the sedolisin family (SedB and SedD). A novel prolyl peptidase, AfuS28, was found to be secreted in both alkaline and acidic conditions. In previous studies, Laps were shown to degrade peptides from their N-terminus until an X-Pro sequence acts as a stop signal. X-Pro sequences can be then removed by DppIV, which allows Laps access to the following residues. We have shown that at acidic pH Seds degrade large peptides from their N-terminus into tripeptides until Pro in P1 or P′1 position acts as a stop for these exopeptidases. However, X-X-Pro and X-X-X-Pro sequences can be removed by AfuS28 thus allowing Seds further sequential proteolysis. In conclusion, both alkaline and acidic sets of proteases contain exoprotease activity capable of cleaving after proline residues that cannot be removed during sequential digestion by nonspecific exopeptidases.
Plant class I glucan endo-1,3-beta-glucosidases (beta-1,3-glucanase; 1,3-beta-D-glucan glucanohydrolase, EC 3.2.1.39) have been implicated in development and defense against pathogen attack. Nevertheless, beta-1,3-glucanase deficiencies generated by antisense transformation of Nicotiana sylvestris and tobacco have little biological effect. We report here that another beta-1,3-glucanase activity is induced in these deficient mutants after infection with necrotizing viruses. Induction of class I beta-1,3-glucanase was markedly inhibited in leaves of N. sylvestris and tobacco antisense transformants infected with tobacco necrosis virus and tobacco mosaic virus, respectively. A serologically distinct beta-1,3-glucanase activity was present in the infected antisense transformants but was absent in both healthy and infected control plants and in antisense transformants treated with the stress hormone ethylene. Immunoblot analyses, localization studies, and measurements of antibody specificity indicate that this compensatory beta-1,3-glucanase activity is an intracellular enzyme different from known tobacco beta-1,3-glucanases. Therefore, plants can compensate for a deficiency in enzyme activity by producing a functionally equivalent replacement--i.e., "ersatz"--protein or proteins. The fact that compensation for beta-1,3-glucanase activity occurs in response to infection argues strongly for an important role of these enzymes in pathogenesis.
