Identifying peptides and sites involved in FtsZ self-assembly in Escherichia coli through in vivo protein photocrosslinking of FtsZ residues mediated by the unnatural amino acid Bpa (p-benzoyl-L-phenylalanine), followed by tandem mass spectrometry analysis of purified FtsZ crosslinking dimers.

ABSTRACT: Bacterial cell division hinges on the Z-ring, an architecture built from the dynamical assembly and disassembly of FtsZ proteins. This delicate balance ensures not only apparent stability, but also continuous remodeling, both of which are required for Z-ring functioning. However, the molecular nature of such subcellular structures remains elusive. In searching for elusive FtsZ self-assembly interfaces involved in assembling the dynamic Z-ring in bacterial cells, we performed in vivo protein photocrosslinking for FtsZ residues as mediated by the unnatural amino acid Bpa and found some N-IDR and N-domain residues were involved in FtsZ self-assembly interactions. To identify counterpart surfaces with which the N-IDR and N-domain interacted when FtsZ self-assembled, we characterized FtsZ peptides and residues that were crosslinked to Bpa residues introduced into these regions by performing tandem mass spectrometry on purified FtsZ crosslinking dimers. Surprisingly, we observed that Bpa residues introduced to the N-IDR (e.g., at T8 or D10) crosslinked with residues in the N-domain, while Bpa residues introduced to the N-domain (e.g., at F40, K51, Q56 or S62) crosslinked with residues in the N-IDR. These crosslinking results strongly suggested that the N-IDR of one subunit interacted with the N-domain of a neighbouring subunit and vice versa when FtsZ self-assembled in cells.

INSTRUMENT(S):

ORGANISM(S): Escherichia Coli

SUBMITTER: Huijia Yin  

LAB HEAD: Zengyi Chang

PROVIDER: PXD063736 | Pride | 2025-05-10

REPOSITORIES: Pride