The transmembrane domain of the T4SS coupling protein TrwB and its role in protein–protein interactions

• Published in: Biochimica et biophysica acta 2013-09, Vol.1828 (9), p.2015-2025
• Main Authors: Segura, Rosa L., Águila-Arcos, Sandra, Ugarte-Uribe, Begoña, Vecino, Ana J., de la Cruz, Fernando, Goñi, Félix M., Alkorta, Itziar
• Format: Article
• Language: English
• Subjects: Bacterial conjugation; Cell Membrane - chemistry; Cell Membrane - metabolism; Conjugation, Genetic - genetics; Conjugative plasmid; Coupling protein; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Escherichia coli - chemistry; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Mutation; Plasmids; Protein Binding; Protein Interaction Domains and Motifs; Protein Structure, Secondary; Protein Transport; Type IV secretion system; VirB10-like proteins
• ISSN: 0005-2736; 0006-3002; 1879-2642
• DOI: 10.1016/j.bbamem.2013.05.022

Bacteria use type IV secretion systems to transfer genetic material and proteins from donor to recipient cells, using proteins encoded by conjugative plasmids. Among those proteins the so-called Type IV Coupling Protein plays a central role in the process. One of the best studied members of this family is TrwB, the conjugative coupling protein of R388 plasmid. Previous studies indicated that the transmembrane domain of TrwB plays a role beyond the mere anchoring of the protein to the membrane. TrwB has also been shown to interact with other conjugative proteins, such as the VirB10-like protein of R388 TrwE. The goal of this study is to elucidate the role of the different domains of TrwB and TrwE in their biological function, and in both self- and TrwB–TrwE interactions. To this aim, a series of TrwB and TrwE deletion mutant proteins were constructed. Conjugation and interaction studies revealed that the transmembrane domain of TrwB, and particularly its second transmembrane helix, is needed for TrwB self-interaction and for R388 conjugative transfer and that there are contacts between TrwB and TrwE in the membrane. On the contrary, the lack of the TMD of TrwE does not completely abolish R388 conjugation although the interaction between TrwE–TrwB is lost. These results identify protein–protein interactions inside the membrane needed for T4SS function. [Display omitted] •The transmembrane domain of TrwB is essential for its biological activity.•The transmembrane domain of TrwB is the minimal element necessary for self-interactions.•TrwE is anchored to inner membrane and its C-terminal is necessary for conjugation.•The transmembrane domains of TrwB and TrwE are involved in TrwB–TrwE interactions.