Two-step fast protein liquid chromatographic purification of the Serratia marcescens hemolysin and peptide mapping with mass spectrometry |
| |
Authors: | Ralf Hertle Roderich Süssmuth Volkmar Braun Günther Jung |
| |
Abstract: | The pore forming toxin of Serratia marcescens (ShlA) is secreted and activated by an outer membrane protein (ShlB). Activation of inactive ShlA (termed ShlA*) by ShlB is dependent on phosphatidylethanolamine (PE). Activation may be a covalent modification of ShlA. To test this hypothesis, the responsible activation domain (in the N-terminal 255 amino acids of ShlA) was isolated from whole bacteria with 8 M urea in an inactive form (ShlA-255*) and from the culture supernatant in an active form (ShlA-255), followed by a two-step purification by anion-exchange chromatography and gel permeation chromatography. Comparison of a tryptic peptide map of both forms with subsequent electrospray mass spectrometry (ES-MS) and sequencing by tandem ES-MS revealed no modification. These data imply that ShlB presumably imposes a conformation on ShlA-255 that triggers activity. |
| |
Keywords: | Hemolysin |
本文献已被 ScienceDirect 等数据库收录! |