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Regulation of multispanning membrane protein topology via post-translational annealing.

Van Lehn RC, Zhang B, Miller TF - Elife (2015)

Bottom Line: This work reveals a mechanism for regulating membrane-protein topogenesis, in which initially misintegrated configurations of the proteins undergo post-translational annealing to reach fully integrated multispanning topologies.The energetic barriers associated with this post-translational annealing process enforce kinetic pathways that dictate the topology of the fully integrated proteins.The proposed mechanism agrees well with the experimentally observed features of EmrE topogenesis and provides a range of experimentally testable predictions regarding the effect of translocon mutations on membrane protein topogenesis.

View Article: PubMed Central - PubMed

Affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, United States.

ABSTRACT
The canonical mechanism for multispanning membrane protein topogenesis suggests that protein topology is established during cotranslational membrane integration. However, this mechanism is inconsistent with the behavior of EmrE, a dual-topology protein for which the mutation of positively charged loop residues, even close to the C-terminus, leads to dramatic shifts in its topology. We use coarse-grained simulations to investigate the Sec-facilitated membrane integration of EmrE and its mutants on realistic biological timescales. This work reveals a mechanism for regulating membrane-protein topogenesis, in which initially misintegrated configurations of the proteins undergo post-translational annealing to reach fully integrated multispanning topologies. The energetic barriers associated with this post-translational annealing process enforce kinetic pathways that dictate the topology of the fully integrated proteins. The proposed mechanism agrees well with the experimentally observed features of EmrE topogenesis and provides a range of experimentally testable predictions regarding the effect of translocon mutations on membrane protein topogenesis.

No MeSH data available.


Simulation snapshot illustrating the initial configuration comprised of 9 CG beads.Positions of several beads in the ribosome (brown) and translocon (green) are labeled, with each labeled bead indicated by a black dot. Positions are labeled in units of σ.DOI:http://dx.doi.org/10.7554/eLife.08697.005
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fig1s2: Simulation snapshot illustrating the initial configuration comprised of 9 CG beads.Positions of several beads in the ribosome (brown) and translocon (green) are labeled, with each labeled bead indicated by a black dot. Positions are labeled in units of σ.DOI:http://dx.doi.org/10.7554/eLife.08697.005

Mentions: Simulations are initialized from equilibrated configurations of the nascent protein, initially comprised of 9 CG beads, with the C-terminus attached to the ribosome exit channel (Figure 1—figure supplement 2). Translation is performed by adding a new CG bead to the C-terminus of the nascent protein and attaching it to the ribosome exit channel; the previous C-terminus is released from the exit channel. The simulation is then continued for 125 ms before the next bead is added, a simulation time which corresponds to a translation rate of 24 residues/s (Bilgin et al., 1992). At the end of translation, the C-terminus is released from the ribosome exit channel and simulations are continued until all beads in the TMDs are at least 4.5σ from the origin and integrated with either a Ncyto/Ccyto or Nperi/Cperi topology. The ribosome remains bound to the translocon for the duration of all simulations (Potter and Nicchitta, 2002; Schaltetzky and Rapoport, 2006). The distance threshold ensures that the final configuration of the protein has exited from both the ribosome and translocon channel.


Regulation of multispanning membrane protein topology via post-translational annealing.

Van Lehn RC, Zhang B, Miller TF - Elife (2015)

Simulation snapshot illustrating the initial configuration comprised of 9 CG beads.Positions of several beads in the ribosome (brown) and translocon (green) are labeled, with each labeled bead indicated by a black dot. Positions are labeled in units of σ.DOI:http://dx.doi.org/10.7554/eLife.08697.005
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4635508&req=5

fig1s2: Simulation snapshot illustrating the initial configuration comprised of 9 CG beads.Positions of several beads in the ribosome (brown) and translocon (green) are labeled, with each labeled bead indicated by a black dot. Positions are labeled in units of σ.DOI:http://dx.doi.org/10.7554/eLife.08697.005
Mentions: Simulations are initialized from equilibrated configurations of the nascent protein, initially comprised of 9 CG beads, with the C-terminus attached to the ribosome exit channel (Figure 1—figure supplement 2). Translation is performed by adding a new CG bead to the C-terminus of the nascent protein and attaching it to the ribosome exit channel; the previous C-terminus is released from the exit channel. The simulation is then continued for 125 ms before the next bead is added, a simulation time which corresponds to a translation rate of 24 residues/s (Bilgin et al., 1992). At the end of translation, the C-terminus is released from the ribosome exit channel and simulations are continued until all beads in the TMDs are at least 4.5σ from the origin and integrated with either a Ncyto/Ccyto or Nperi/Cperi topology. The ribosome remains bound to the translocon for the duration of all simulations (Potter and Nicchitta, 2002; Schaltetzky and Rapoport, 2006). The distance threshold ensures that the final configuration of the protein has exited from both the ribosome and translocon channel.

Bottom Line: This work reveals a mechanism for regulating membrane-protein topogenesis, in which initially misintegrated configurations of the proteins undergo post-translational annealing to reach fully integrated multispanning topologies.The energetic barriers associated with this post-translational annealing process enforce kinetic pathways that dictate the topology of the fully integrated proteins.The proposed mechanism agrees well with the experimentally observed features of EmrE topogenesis and provides a range of experimentally testable predictions regarding the effect of translocon mutations on membrane protein topogenesis.

View Article: PubMed Central - PubMed

Affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, United States.

ABSTRACT
The canonical mechanism for multispanning membrane protein topogenesis suggests that protein topology is established during cotranslational membrane integration. However, this mechanism is inconsistent with the behavior of EmrE, a dual-topology protein for which the mutation of positively charged loop residues, even close to the C-terminus, leads to dramatic shifts in its topology. We use coarse-grained simulations to investigate the Sec-facilitated membrane integration of EmrE and its mutants on realistic biological timescales. This work reveals a mechanism for regulating membrane-protein topogenesis, in which initially misintegrated configurations of the proteins undergo post-translational annealing to reach fully integrated multispanning topologies. The energetic barriers associated with this post-translational annealing process enforce kinetic pathways that dictate the topology of the fully integrated proteins. The proposed mechanism agrees well with the experimentally observed features of EmrE topogenesis and provides a range of experimentally testable predictions regarding the effect of translocon mutations on membrane protein topogenesis.

No MeSH data available.