Plasticity within the barrel domain of BamA mediates a hybrid-barrel mechanism by BAM

AbstractIn Gram-negative bacteria, the biogenesis of β-barrel outer membrane proteins is mediated by the β-barrel assembly machinery (BAM). The mechanism employed by BAM is complex and so far- incompletely understood. Here, we report the structures of BAM in nanodiscs, prepared using polar lipids and native membranes, where we observe an outward-open state. Mutations in the barrel domain of BamA reveal that plasticity in BAM is essential, particularly along the lateral seam of the barrel domain, which is further supported by molecular dynamics simulations that show conformational dynamics in BAM are modulated by the accessory proteins. We also report the structure of BAM in complex with EspP, which reveals an early folding intermediate where EspP threads from the underside of BAM and incorporates into the barrel domain of BamA, supporting a hybrid-barrel budding mechanism in which the substrate is folded into the membrane sequentially rather than as a single unit.

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The membranes of Gram-negative bacteria: progress in molecular modelling and simulation

Biochemical Society Transactions ◽

10.1042/bst20140262 ◽

2015 ◽

Vol 43 (2) ◽

pp. 162-167 ◽

Cited By ~ 15

Author(s):

Syma Khalid ◽

Nils A. Berglund ◽

Daniel A. Holdbrook ◽

Yuk M. Leung ◽

Jamie Parkin

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Lipid Bilayers ◽

Molecular Modelling ◽

Gram Negative Bacteria ◽

Gram Negative ◽

Bacterial Membranes ◽

Lipid Species ◽

Membrane Models ◽

Dynamics Simulations

Molecular modelling and simulations have been employed to study the membranes of Gram-negative bacteria for over 20 years. Proteins native to these membranes, as well as antimicrobial peptides and drug molecules have been studied using molecular dynamics simulations in simple models of membranes, usually only comprising one lipid species. Thus, traditionally, the simulations have reflected the majority of in vitro membrane experimental setups, enabling observations from the latter to be rationalized at the molecular level. In the last few years, the sophistication and complexity of membrane models have improved considerably, such that the heterogeneity of the lipid and protein composition of the membranes can now be considered both at the atomistic and coarse-grain levels of granularity. Importantly this means relevant biology is now being retained in the models, thereby linking the in silico and in vivo scenarios. We discuss recent progress in simulations of proteins in simple lipid bilayers, more complex membrane models and finally describe some efforts to overcome timescale limitations of atomistic molecular dynamics simulations of bacterial membranes.

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Glucose transport via the pseudomonad porin OprB: implications for the design of Trojan Horse anti-infectives

Physical Chemistry Chemical Physics ◽

10.1039/c9cp00778d ◽

2019 ◽

Vol 21 (16) ◽

pp. 8457-8463 ◽

Cited By ~ 2

Author(s):

Joan Coines ◽

Silvia Acosta-Gutierrez ◽

Igor Bodrenko ◽

Carme Rovira ◽

Matteo Ceccarelli

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Glucose Transport ◽

Trojan Horse ◽

Gram Negative Bacteria ◽

Gram Negative ◽

Dynamical Features ◽

Dynamics Simulations

Knowing the structural and dynamical features of specific porins from poor-permeable Gram-negative bacteria helps to design anti-infectives with optimal permeation. Molecular dynamics simulations can characterize and quantify the transport of substrates through these specific porins.

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Molecular Basis of Glucose Transport Mechanism in Plants

10.26434/chemrxiv.8049848.v1 ◽

2019 ◽

Cited By ~ 2

Author(s):

Balaji Selvam ◽

Ya-Chi Yu ◽

Liqing Chen ◽

Diwakar Shukla

Keyword(s):

Molecular Dynamics ◽

Free Energy ◽

Molecular Dynamics Simulations ◽

Conformational Changes ◽

Transport Mechanism ◽

Conformational Dynamics ◽

Site Directed Mutagenesis ◽

Free Energy Barrier ◽

Transport Cycle ◽

Dynamics Simulations

<p>The SWEET family belongs to a class of transporters in plants that undergoes large conformational changes to facilitate transport of sugar molecules across the cell membrane. However, the structures of their functionally relevant conformational states in the transport cycle have not been reported. In this study, we have characterized the conformational dynamics and complete transport cycle of glucose in OsSWEET2b transporter using extensive molecular dynamics simulations. Using Markov state models, we estimated the free energy barrier associated with different states as well as 1 for the glucose the transport mechanism. SWEETs undergoes structural transition to outward-facing (OF), Occluded (OC) and inward-facing (IF) and strongly support alternate access transport mechanism. The glucose diffuses freely from outside to inside the cell without causing major conformational changes which means that the conformations of glucose unbound and bound snapshots are exactly same for OF, OC and IF states. We identified a network of hydrophobic core residues at the center of the transporter that restricts the glucose entry to the cytoplasmic side and act as an intracellular hydrophobic gate. The mechanistic predictions from molecular dynamics simulations are validated using site-directed mutagenesis experiments. Our simulation also revealed hourglass like intermediate states making the pore radius narrower at the center. This work provides new fundamental insights into how substrate-transporter interactions actively change the free energy landscape of the transport cycle to facilitate enhanced transport activity.</p>

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