Fig 5.
Phosphopantetheinylation of IacP by AcpS. (A) Analysis by MALDI-TOF mass spectrometry of the intact IacP_CBP protein recovered after TAP when purified from strain PBADacpS IacP_TAP (JV68), which was grown while AcpS was produced (0.1% arabinose; black spectrum) or depleted (0.1% glucose; gray spectrum). The lower molecular mass (14,250.4 Da) indicated for the major gray peak corresponds to the global mass of apo-IacP_CBP ([M + H]+ theoretical mass of 14,252.2 Da for apo-IacP_CBP), while the higher molecular mass (14,590 Da) indicated for the major black peak corresponds to the global mass of holo-IacP_CBP ([M + H]+ theoretical mass of 14,591.5 Da for holo-IacP_CBP). (B) Western blot on protein crude extracts using PAP to reveal the ProtA part of TAP-tagged IacP. Excepted if specified (+ Ara 0.1%), strains were grown under noninducing conditions of PBAD, meaning that AcpS production from the chromosome was off when we used the strain PBADacpS IacP_TAP (JV68). When specified, strains harbored a plasmid carrying one of the PPTases under the control of PTET.