NMR characterization of the Type III Secretion System Tip Chaperone Protein PcrG of Pseudomonas aeruginosa

Abstract

Lung infection with Pseudomonas aeruginosa is the leading cause of death among cystic fibrosis patients. To initiate infection, P. aeruginosa assembles a protein nanomachine, the type III secretion system (T3SS) to inject bacterial proteins directly into target host cells. An important regulator of the P. aeruginosa T3SS is the chaperone protein PcrG, which forms a complex with the tip protein, PcrV. In addition to its role as a chaperone to the tip protein, PcrG also regulates protein secretion. PcrG homologs are also important in the T3SS of other pathogens such as Yersinia pestis, the causative agent of bubonic plague. The atomic structure of PcrG or any member of the family of tip protein chaperones is currently unknown. Here, we show by CD and NMR spectroscopy that PcrG lacks a tertiary structure. However, it is not completely disordered but contains secondary structures dominated by two long α-helices from residues 16–41 and 55–76. NMR backbone dynamics data show that the helices in PcrG have semi-rigid flexibility and they tumble as a single entity with similar backbone dynamics. NMR titrations show that the entire length of PcrG residues from 9–76 is involved in binding to PcrV. Thus the PcrG family of T3SS chaperone proteins is essentially partially folded.