crystal structure of prgi-sipd insight into a secretion competent state of the type three secretion system needle tip and its interaction with host ligands晶体结构的prgi-sipd洞察三型分泌系统的分泌能力状态针尖端与主机配体及其交互.pdfVIP

Crystal Structure of PrgI-SipD: Insight into a Secretion Competent State of the Type Three Secretion System Needle Tip and its Interaction with Host Ligands 1 2 1 1 Michele Lunelli , Robert Hurwitz , Jutta Lambers , Michael Kolbe * 1 Max Planck Institute for Infection Biology, Department of Cellular Microbiology, Berlin, Germany, 2 Max Planck Institute for Infection Biology, Core Facility Protein Purification, Berlin, Germany Abstract Many infectious Gram-negative bacteria, including Salmonella typhimurium, require a Type Three Secretion System (T3SS) to translocate virulence factors into host cells. The T3SS consists of a membrane protein complex and an extracellular needle together that form a continuous channel. Regulated secretion of virulence factors requires the presence of SipD at the T3SS needle tip in S. typhimurium. Here we report three-dimensional structures of individual SipD, SipD in fusion with the needle subunit PrgI, and of SipD:PrgI in complex with the bile salt, deoxycholate. Assembly of the complex involves major conformational changes in both SipD and PrgI. This rearrangement is mediated via a p bulge in the central SipD helix and is stabilized by conserved amino acids that may allow for specificity in the assembly and composition of the tip proteins. Five copies each of the needle subunit PrgI and SipD form the T3SS needle tip complex. Using surface plasmon resonance spectroscopy and crystal structure analysis we found that the T3SS needle tip complex binds deoxycholate with micromolar affinity via a cleft formed at the SipD:PrgI interface. In the structure-based three-dimensional model of the T3SS needle tip, the bound deoxycholate faces the host membrane. Recently, binding of SipD with bile salts present in the