Differential effect of site-directed mutations in pelC on pectate lyase activity, plant tissue maceration, and elicitor activity.

Oligonucleotide site-directed mutations were introduced into the pelC gene of Erwinia chrysanthemi EC16 that directed single or double amino acid changes affecting disulfide linkages, calcium binding, catalysis, and protein folding. Subsequent characterization of the purified PelC mutant proteins demonstrated that pectinolytic function involves amino acids located near the calcium binding site rather than those surrounding an invariant vWiDH sequence. Wild-type PelC and the tested mutant proteins generally macerated plant tissue in proportion to their specific pectinolytic activity in vitro. However, some mutants gave higher maceration activity in plant tissue and elicited greater production of the phytoalexin, glyceollin, in soybean cotyledons than predicted by their in vitro pectinolytic activity. Most notable in this regard were three different mutations at lysine 172 with greatly reduced pectinolytic activity but as much elicitor activity as the wild-type protein. PelE macerated plant tissue 10 times more efficiently than PelC, as observed previously, but surprisingly showed equal activity in the elicitor assay. The results indicate that factors other than pectinolytic activity per se are involved in plant tissue maceration and elicitor activity.