Direct observation of the uptake of outer membrane proteins by the periplasmic chaperone Skp.
The transportation of membrane proteins through the aqueous subcellular space is an important and challenging process. Its molecular mechanism and the associated structural change are poorly understood. Periplasmic chaperones, such as Skp in Escherichia coli, play key roles in the transportation and protection of outer membrane proteins (OMPs) in Gram-negative bacteria. The molecular mechanism through which Skp interacts with and protects OMPs remains mysterious. Here, a combined experimental and molecular dynamics simulation study was performed to gain the structural and dynamical information in the process of OMPs and Skp binding. Stopped-flow experiments on site specific mutated and labeled Skp and several OMPs, namely OmpC, the transmembrane domain of OmpA, and OmpF, allowed us to obtain the mechanism of OMP entering the Skp cavity, and molecular dynamics simulations yielded detailed molecular interactions responsible for this process. Both experiment and simulation show that the entrance of OMP into Skp is a highly directional process, which is initiated by the interaction between the N-terminus of OMP and the bottom "tentacle" domain of Skp. The opening of the more flexible tentacle of Skp, the non-specific electrostatic interactions between OMP and Skp, and the constant formation and breaking of salt bridges between Skp and its substrate together allow OMP to enter Skp and gradually "climb" into the Skp cavity in the absence of an external energy supply.
State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Department of Chemical Biology, Biodynamic Optical Imaging Center, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
Zhi-Xin
ZX
Lyu
Qiang
Q
Shao
Yi Qin
YQ
Gao
Xin Sheng
XS
Zhao
D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism*; D002942:Circular Dichroism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D008962:Models, Theoretical; D018832:Molecular Chaperones / Q000378:metabolism*; D019897:Periplasm / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
D001425:Bacterial Outer Membrane Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; C072721:Skp protein, E coli