Reconstitution of a nanomachine driving the assembly of proteins into bacterial outer membranes

• DOI: 10.1038/ncomms6078
• Publication Date: 2014-10-24T10:48:16Z
• Authors: Hsin-Hui Shen; Denisse L. Leyton; Takuya Shiota; Matthew J. Belousoff; Nicholas Noinaj; Jingxiong Lu; Stephen A. Holt; Khershing Tan; Joel Selkrig; Chaille T. Webb; Susan K. Buchanan; Lisandra L. Martin; Trevor Lithgow
• Keywords: Quartz Crystal Microbalance; Component (thermodynamics)

In biological membranes, various protein secretion devices function as nanomachines, and measuring the internal movements of their component parts is a major technological challenge. The translocation and assembly module (TAM) is a nanomachine required for virulence of bacterial pathogens. We have reconstituted a membrane containing the TAM onto a gold surface for characterization by quartz crystal microbalance with dissipation (QCM-D) and magnetic contrast neutron reflectrometry (MCNR). The MCNR studies provided structural resolution down to 1 Å, enabling accurate measurement of protein domains projecting from the membrane layer. Here we show that dynamic movements within the TamA component of the TAM are initiated in the presence of a substrate protein, Ag43, and that these movements recapitulate an initial stage in membrane protein assembly. The reconstituted system provides a powerful new means to study molecular movements in biological membranes, and the technology is widely applicable to studying the dynamics of diverse cellular nanomachines.