Figure 5
Disulfide-pairing determination of the major oxidative folding intermediates of NvCI. CM stands for carboximethyl cysteine (alkylated with IAA) and PE stands for pyridylethyl cysteine (alkylated with VP). The cysteine positions are indicated above the graph. (a) Characterization of fraction I: The purified fraction I was derivatized with IAA, reduced with DTT and further derivatized with VP prior trypsin digestion. The molecular masses determined by MALDI-TOF-MS for the derivatized tryptic peptides are shown. The 1715.8 Da derivatized peptide was sequenced by MALDI-TOF-MS2 and the spectrum characterized by β and γ ion series with the assigned sequence is shown. (b) Fraction IIb characterization: The purified IIab fraction was derivatized with VP and purified by RP-HPLC (see Fig. 5c, II b-PE). Then it was reduced with DTT, derivatized with IAA and subjected to trypsin digestion and MALDI-TOF analysis. The 1715.8 Da fragment was sequenced by MALDI-TOF-MS2 and the spectrum with the assigned sequence is shown. (c) Fraction IIa characterization: The purified IIab fraction was derivatized with IAA and purified by RP-HPLC (see Fig. 5, II a-CM). Then it was reduced with DTT, derivatized with VP and subjected to trypsin digestion and MALDI-TOF analysis. The 1763.8 Da fragment was sequenced by MALDI-TOF-MS2 and the assigned sequence is shown.
