Figure 2.

Fraction and identification of the active component in pectolyase that can restore reconstituted expansin-induced extension in mature cell walls. A, Fractionations of pectolyase from a CM-Sepharose column showed a single peak (peak II) of activity (solid line, the absorbance; dashed line, the restoring activity). B, Fractionations of peak II in A from Bio-Gel P-60 gel-permeation chromatography showed two overlapping peaks (1 and 2) of activity (solid line, the absorbance; dashed line, the restoring activity). C, Silver-stained SDS-PAGE of proteins in collecting tubes around peaks 1 and 2, in which five protein bands were, respectively, marked as a, b, c, d, and e. D, Peptide sequences of proteins b and d identified by LC-ESI-MS/MS. Protein bands b and d were excised from the gel and digested with trypsin. The digested peptides were subjected to LC-ESI-MS/MS analysis using a Finnigan LCQ Deca XP Plus LC/MS/MS. Identification was obtained by comparing the experimental data with the NCBI nonredundant protein database and was validated after considering at least four peptide sequences per protein (Bruneel et al., 2005).