RELEASE OF CONTENT THROUGH MECHANO- SENSITIVE GATES IN PRESSURISED LIPOSOMES. Martti Louhivuori University of Groningen
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1 RELEASE OF CONTENT THROUGH MECHANO- SENSITIVE GATES IN PRESSURISED LIPOSOMES Martti Louhivuori University of Groningen
2 MARTINI coarse-grained model
3 water P4 butane Qo Na DPPC cholesterol peptide!alywk" Qa SP1 C1 C5 SC3 SP1 SNd SC1 SC4 C1 C1 Qd SC4 SC4 C3 No MARTINI CG model interaction sites
4 parametrisation of MARTINI experimental!thermodynamic" data! non#bonded interactions atomistic MD simulations! bonded interactions
5 distribution angle parametrisation bonded interactions
6 parametrisation non#bonded interactions
7 THE VALIDATION comparing to experimental measurements Rhombohedral phase (experimentally observed for DOPC/DOPE 3:1 and 2:1 Lyan & Huang, 2002) side view top view Reproduced in CG simulation (Marrink & Mark, Biophys. J., 2004)
8 bilayers rafts vesicles sugars vesicles w/ proteins membrane proteins
9 bilayers rafts vesicles sugars vesicles w/ proteins membrane proteins
10 water P4 butane SC4 Qo Na DPPC cholesterol peptide!alywk" SPEED Qa short#range interactions large time#step few degrees of freedom EASE of USE C1 building#block approach limited # of particles physical units SP1 SC3 SC1 C1 GENERAL C1 consistent modeling biomolecular systems SP1 easily extended SC4 ACCURACY parametrisation based on Qd thermodynamic data C3 multi#level optimisation C5 SNd SC4 No MARTINI CG model interaction sites
11 mmm! ouch! drugs patient targeted drug release. how?
12 drug delivery vehicle nanobot BOOM
13 drug delivery vehicle nanobot
14 mechano-sensitive channels safety valves of cell sense tension in the membrane MscL, MscK, MscS, MscM < 10 mn/m PDB: 2VV5
15 top bottom MscL controllable activation & non!selective conductance
16 MscL activity flickering conductivity multiple levels! subconductive states activation < 1 ms de#activation 1$100 ms Sukharev et al.!1997" Annu Rev Physiol 59: 633$657
17 non-selective channel no ion selectivity even small proteins pass through! 15$20 Å Cruickshank et al.!1997" Biophys J 73: 1925$1931
18 photosensitivivity attached compound undergoes light induced charge separation reversible localised Koçer et al.!2005" Science 309: 755$758
19 photosensitivivity photosensitive lipids used to transfer signal to mechanical stress reversible localised Folgering et al.!2004" Langmuir 20: 6985$6987
20 nano-container aka liposome nano-particles aka drugs
21 nano-transporter liposomes tiny lipid vesicles membrane fusion trans#membrane transport drug delivery curvature e%ects MscL mechano#sensitive pressure valves of cells touch & hear non#selective, large membrane channel
22 game plan BOOM
23 MscL saves the day
24 660 kn/m s! lysis 140 kn/m s! ok oh-oh!
25 analysis
26 analysis closed open 5 us
27 activation mechanism
28 postactivation 67 mn/m " 24 nm 1.04 H2O / ns
29 equilibrium? tension pressure tension (mn/m) / pressure (bar) (-1.12 ± 0.05) bar/µs 20 (-0.54 ± 0.02) mn m /µs (93 ± 4) µs (86 ± 4) µs time (µs)
30 MFFA boundary potentials RDF r [nm] mimic interactions with bulk solvent 30 MFFA Risselada et al.!2008" J Phys Chem B 112: 7438$ r [nm]
31 pumping water into liposomes additional mean#field potential inside the liposome start with r = 0.01 nm increase slowly for 20ns until r = 3.9 nm fill the cavity with water, relax and repeat as needed
32 water-repellant lipid tails modified Lennard# Jones potential against water DOPC NC3 PO4 wdopc NC3 PO4 C5 wc5 epsilon sigma [ (σ ) 12 ( ) ] σ 6 V (r) =4ɛ r r GL1 C1 C2 D3 C4 C5 GL2 C1 C2 D3 C4 C5 C1 C2 D3 C4 wc5 GL1 GL2 C1 C2 D3 C4 wc5
33 Ollila et al.!2009" Phys Rev Lett, 102: D pressure fields divide system into a 3D grid use local virial for each volume element calculate averages P local (r) = 1 V i δ (r r i ) m i v i v i i j F ij C i j δ(r 1)dl
34 flow rate Partial densities 2000 Density (kg m -3 ) R! Protein wdopc W PO4! R Box (nm)
35 summary large#scale biological systems accessible to CG simulations release of an osmotic shock via MscL activation achieved water flux OUT:!6.0 ± 1.3"& ions/ns IN:!1.7 ± 0.3"& ions/ns MODEL: 0.2$40& ions/ns & Steinbacher et al.!2007" & CurrTopicsMembranes 58:1#24 pore radius!11.6 ± 0.8" Å MscL activation is indeed a last#ditch e%ort to prevent lysis iris#like, non#symmetric opening &!exp. 15$20 Å" blocking of the channel by the cytoplasmic helices a first step in closure?
36 future dye molecules to directly compare our release of nano#particles to experimental data nano#pores formed by other molecules, e.g. anti#microbial peptides activation of the channel using lyso#lipids
37 Siewert#Jan Marrink Erik van der Giessen Jelger Risselada UPPSALA UNIVERSITET David van der Spoel TAMPERE UNIVERSITY OF TECHNOLOGY Samuli Ollila Ilpo Vattulainen Stockholm University Erik Lindahl
38
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