Computer Simulation of Membrane Proteins: Membrane Deformation, Signal Transduction and Cellular Uptake of NanoparticlesTongtao Yue and Xianren Zhang Division of Molecular and Materials Simulation, State Key Laboratory of Organic-Inorganic CompositesBeijing University of Chemical TechnologyBeijing 100029, China

Clustering of membrane anchored proteins and its role in membrane deformation and signal transductionLipid membraneReceptor-mediated interaction between membrane and nanoparticles with different properties

Story 1:The Relationship Between Membrane Deformation and Clustering of Anchored ProteinsMembrane curvature is ubiquitous in cell environmentHow to deform a membrane?Mcmahon et al, Nature, 438, 590 (2005)

What if the clustering of anchored proteins is taken into account?Mcmahon et al, Nature, 438, 590 (2005)

The different protein hydrophobic lengths not only result in different aggregation numbers, but also lead to different aggregation dynamics.Protein Clustering:

Membrane DeformationEffect of Membrane Surface Tension:Membrane deformation by the anchored proteins of ntp =6, at different values of lipid density: (A) 1.47; (B) 1.67; (C) 1.69; and (D) 1.73. The figure shows typical snapshots (left), the corresponding best-fitted geometric surfaces (bottom right) and curvature distribution (top right).A negative membrane tension is a prerequisite for the bending of a lipid bilayer, and the deformation of the membrane is found to be located mainly around the protein clusters.T. Yue et al, Soft Matter, 2010, 6, 6109

Membrane Deformation Effect of Protein Hydrophobic Length:Membrane deformation by the different anchored proteins, (A) ntp =3; (B) ntp =4; (C) ntp =6; and (D) ntp =7. (E) shows the the largest membrane curvature as a function of lipid density.Deep penetration of proteins into membrane would result into their clustering, which in turn produces positive curvature considerably exceeding that for shallow inserting proteins

Membrane Deformation Effect of Protein Hydrophobic Length:(a) The largest membrane curvature as a function of LNPA, and (b) the largest membrane curvature as a function of the hydrophobic length of anchored proteins. Note that LNPA is set to 1.70 in (b).T. Yue et al, Soft Matter, 2010, 6, 6109

Membrane Vesiculation:Typical snapshots during the formation of a vesicle.Before observable membrane curvature appears, the clustering of proteins occurs because of the strong effective attraction between proteins due to theirperturbations on the membrane

Protein clustering Senses Membrane Curvature:Distribution of the cluster size (top) and corresponding local curvature (bottom) as a function of cluster sizeMost protein clusters are located on membrane positions with positive curvature. More importantly, larger protein clusters tend to be located atmembrane positions with higher positive curvature.T. Yue et al, Soft Matter, 2010, 6, 6109

123Summaries:Curvature production is enhanced by the clustering of anchored proteins, and the enhancement depends on the protein hydrophobic length.For the membrane proteins with deep insertion, the clustering of proteins may induce membrane vesiculation at negative membrane tensions.The protein clustering can sense the membrane curvature, although the way they respond to the local curvature again depends on the protein hydrophobic length.T. Yue et al, Soft Matter, 2010, 6, 6109

Story 2:Signal Transduction Across Cellular Membrane can be Mediated by Coupling of the Clustering of Anchored Proteins in Both LeafletsOne of the key questions regarding signal transduction is how the signal received by outer-leaflet is relayed to the inner-leaflet.How is the coupling occurs between anchored proteins in different leaflet?Kusumi et al, Traffic (2004)

Three Coupling Patterns of Protein Clustering:(a-c) Snapshots for three coupling patterns of protein clustering. (a) A face-to-face clustering (top: n=3; bottom: n=6), (b) an interdigitated clustering (top: n=6; bottom: n=6), and (c) a weak-coupled clustering (top: n=4; bottom: n=4). Blue represents proteins in the bottom leaflet and pink represents proteins in the upper leaflet. (d) shows two-dimensional radial distribution function (RDF) of proteins in one leaflet with respective to those in the opposite leaflet.The coupling pattern are strongly dependent on the hydrophobic length of proteins in both leaflets.T. Yue and X. Zhang, Phys. Rev. E, 2012, 85, 011917

Movies:Face-to-face clusteringInterdigitated clusteringT. Yue and X. Zhang, Phys. Rev. E, 2012, 85, 011917

Both upper and bottom leaflets are strongly perturbed by the clustering of anchored proteins in one leaflet. The membrane perturbation induced by upper protein cluster thus tends to cause the proteins in the bottom leaflet to redistribute.Z-coordinates for lipids adjacent to the anchored protein cluster. The local membrane thickness and lipid order parameter are given in the left and right insets. The hydrophobic length of the anchored proteins is set to (a) n=3, (b) n=4, (c) n=5, and (d) n=6.Membrane Perturbation:

Trajectories:T. Yue and X. Zhang, Phys. Rev. E, 2012, 85, 011917

Protein Clustering and Phase Diagram:The extent of protein clustering is found to be affected by the coupling patterns, which depends on the hydrophobic length of proteins in both leaflets: weak coupled clustering: face-to-face clustering: interdigitated clustering

123Summaries:The disturbance of the clustering of anchored proteins in one leaflet can extend across the full thickness of a bilayer, thus inducing proteins in the opposite leaflet to redistribute.Depending on the hydrophobic length of anchored membrane proteins, three coupling patterns are observed.We proposed a new mechanism for signal transduction via coupling of protein clustering in this work, this mechanism shows particular selectivity in the downstream signaling.T. Yue and X. Zhang, Phys. Rev. E, 2012, 85, 011917

Story 3:Molecular Understanding of Receptor-mediated Membrane Responses to Ligand-coated NanoparticlesNanotechnology Present a Janus Face!How to maximize the efficiency of drug delivery while minimize their cytotoxicity?McNerny et al. Nanomed. and Nanobio. (2010)Huajian Gao et al, PNAS (2005)

Receptor mediated endocytosisNanoparticle adhesionNanoparticle penetrationNanoparticle induced membrane ruptureFour Kinds of Membrane Responses:T. Yue and X. Zhang, Soft Matter, 2011, 7, 9104

Receptor Mediated Endocytosis:Time evolution of the extent of wrapping.The endocytosis is controlled by the balance of receptor-ligand binding energy and membrane bending energy.

Receptor Mediated EndocytosisEffect of Ligand Density:Time evolution of the extent of endocytosis for NPs with different ligand densities, NL=18 (black line), and NL=40 (red line).NPs with more coated ligands are more easily wrapped by the membrane.T. Yue and X. Zhang, Soft Matter, 2011, 7, 9104

Adhesion and Penetration:(A) The evolution of the percentage of surface beads of the NP contacting with the membrane and (B) the evolution of the NP-membrane distance.Adhesion and penetration of NPs are mainly determined by NPs size and membrane surface tension. Larger NPs tend to adhere on the membrane with larger membrane tension and vice versa.

NP Induced Membrane Rupture:NPs with smaller size and larger ligand density are found to induce the membrane rupture more easily.T. Yue and X. Zhang, Soft Matter, 2011, 7, 9104

Morphology Diagram:Morphology diagram of the membrane responses to adsorption of NP with 18 ligands coated in the plane of membrane tension and NP radius.

123Summaries:There exist four kinds of membrane responses, including receptor-mediated endocytosis, NP adhesion, NP penetration, and membrane rupture.NP size, ligand density on the NP surface, and membrane surface tension are all crucial in the membrane responding processes.The phase diagrams presented here have the potential to provide both qualitative and quantitative guidelines for the designing of novel drug delivery biomaterials.T. Yue and X. Zhang, Soft Matter, 2011, 7, 9104

Story 4:Cooperative Effect in Receptor-Mediated Endocytosis of Multiple NanoparticlesEndocytosis of Single NPNP sizeNP shapeHow about the endocytosis of multiple NPs?Chithrani et al, Nano Lett. (2007)Reynwar et al, Nature. (2007)

Internalization of Nine Identical NPs:D=2.5nm: NPs generally cluster together on the membrane D=4.0nm: NPs aggregate into pearl-chain-like arrangementsD=6.0nm: Independent endocytosisT. Yue and X. Zhang, ACS Nano, 2012, 6, 3196

Movies:T. Yue and X. Zhang, ACS Nano, 2012, 6, 3196

Movies:T. Yue and X. Zhang, ACS Nano, 2012, 6, 3196

Internalization of Two Identical NPs:Internalization pathway of two identical NPs are strongly dependent on the NPs size. Smaller NPs tend to internalize together, while larger NPs tend to internalize independently.

Internalization of Two NPs with Different Size:One NPs diameter is fixed to be 4.5nm, while the diameter of the other NP is (A) 2.5nm, (B) 3.3nm, (C) 4.0nm. Asynchronous internalizationSynchronous internalizationT. Yue and X. Zhang, ACS Nano, 2012, 6, 3196

Pinocytosis-like Internalization of Two NPs with Different Size :(A) Typical snapshots showing the pinocytosis-like internalization of two NPs with different size. (B) The evolution of wrapping extent for two NPs. (C) evolution of COMs along membrane normal direction and that of distance between two NPs (inset)Pinocytosis-like internalization tends to occur for two NPs with large size difference and intermediate initial distance.

Other Internalization pathway for Two NPs with Different Size :When two NPs with large size difference and longer initial distance are placed on the membrane, only the larger NP is wrapped while the smaller NP keep adhering on the membrane

Morphology Diagram:: Independent internalization pathway : Pinocytosis-like internalization pathway: Asynchronous internalization : Synchronous internalization. Note that the diameter of the larger NP is fixed to 4.5nmThe internalization pathway for two NPs with different size is mainly determined by their size difference and initial distanceT. Yue and X. Zhang, ACS Nano, 2012, 6, 3196

123Summaries:Smaller NPs tend to form clusters while internalization. this cooperative effect is weakened by increase the NPs size.The internalization of two identical NPs confirms the importance of NPs size to their internalization pathway.Four different internalization pathways for two NPs with different size are observed which is mainly determined by their size difference and initial distance.T. Yue and X. Zhang, ACS Nano, 2012, 6, 3196

Story 5:Molecular modeling of the pathways of vesiclemembrane interactionShillcock and Lipowsky, Nature Mater. 2005Yi, Shi, and Gao, Phys. Rev. Lett. 2011Advantages of soft NPsHigh solubilityHigh environmental sensitivityLow toxicity

Time sequence of snapshots corresponding to the vesicle hemi-fusion (A) and fusion (B) respectively. The values of LNPA are fixed to 1.54 (A) and 1.16 (B), respectively.Membrane Fusion and Hemifusion:

Vesicle Adhesion:In this figure the receptor density is fixed to 50% under the condition of aRL =4.0 and LNPA =1.1.

Vesicle Rupture:In this figure the receptor density is fixed to 50% under the condition of aRL =0.0 and LNPA =1.66.

Vesicle Endocytosis:In this figure the receptor density is fixed to 50% under the condition of aRL =4.0 and LNPA =1.68.

Effect of Vesicle Elasticity:The self-adjustment of vesicle shape bypasses the high energy barrier of membrane bending to wrap an oblate vesicle, thus facilitates the receptor-mediated endocytosis.

Phase Diagram:: Vesicle Fusion: Vesicle Hemifusion: Vesicle Adhesion, Rupture, or Endocytosis

Effect of Vesicle-Membrane Adhesion Strength:As the adhesion strength decreases, the pathways for the vesicle-membrane interaction changes from vesicle rupture, vesicle endocytosis to vesicle adhesion.

Effect of Membrane Surface Tension:Strong vesicle-membrane adhesion strengthWeak vesicle-membrane adhesion strengthHigher membrane tension would promote vesicle rupture, while lower membrane tension would facilitate vesicle endocytosis.

Effect of Receptor Density:Strong vesicle-membrane adhesion strengthWeak vesicle-membrane adhesion strengthIncrease of receptor density can accelerate the vesicle rupture at strong vesicle-membrane adhesion strength, while at weak vesicle-membrane adhesion strength, increase of receptor density can facilitate the endocytosis process.

Effect of Vesicle Tension:The vesicle with a lower vesicle tension tends to be wrapped by the membrane more efficiently because the vesicle can deform more easily.

Effect of Ligand Density:Both wrapping rate and extent are determined by the ligand density, and vesicles with lower ligand density can not be fully wrapped by the membrane.

12Summaries:Different vesicle responses to the vesicle-membrane adhesion, including vesicle fusion, vesicle hemi-fusion, vesicle adhesion, vesicle endocytosis and vesicle rupture, are observed from our simulations.We also investigate how the pathways of vesicle-membrane interaction depend on the adhesion strength and the membrane and vesicle properties..Tongtao Yue and Xianren Zhang, Submitted

Thank you for your attention!http://www.ms.buct.edu.cn/tt1202@126.comzhangxr@mail.buct.edu.cn