Membrane mediated phase separation of the bacterial nucleoid occlusion protein Noc.

Liquid-liquid phase separation is a fundamental biophysical process to organize eukaryotic and prokaryotic cytosols. While many biomolecular condensates are formed in the vicinity of, or even on lipid membranes, little is known about the interaction of protein condensates and lipid bilayers. In this study, we characterize the recently unknown phase behavior of the bacterial nucleoid occlusion protein Noc. We find that, similarly to other ParB-like proteins, CTP binding tightly regulates Noc's propensity to phase separate. As CTP-binding and hydrolysis also allows Noc to bind and spread on membranes, we furthermore establish Noc condensates as model system to investigate how lipid membranes can influence protein condensation and vice versa. Last, we show that Noc condensates can recruit FtsZ to the membrane, while this does not happen in the non-phase separated state. These findings suggest a new model of Noc mediated nucleoid occlusion, with membrane-mediated liquid-liquid phase separation as underlying principle of complex formation and regulation thereof.

Discriminatory Interaction Behavior of Lipid Vesicles toward Diversely Emissive Carbon Dots Synthesized from Ortho, Meta, and Para Isomeric Carbon Precursors.

Photoluminescent carbon dots (C-dots) are widely used for bioimaging techniques to study different cellular processes. However, biocompatibility of C-dots is crucial because the wrong selection of C-dots may lead to an adverse effect on a particular cellular process. Herein, we investigate the interaction of zwitterionic lipid vesicles with photoluminescent C-dots derived from different isomeric (ortho, meta, and para) precursors of phenylenediamine (PDA) by spectroscopic and microscopic imaging techniques as well as dynamic light scattering methods. The study reveals that interaction of lipid vesicles with C-dots is highly dependent on the properties of the isomeric precursors. We find that vesicles retain their morphology upon interaction with ortho C-dots (oCD). The microscopic images reveal that oCD are selectively embedded in the lipid vesicles and can effectively be used for imaging purpose. On the contrary, meta and para C-dots (mCD and pCD) being located on the interfacial region induce aggregation in the vesicles. We explain the observation in terms of the location of the C-dots on the lipid vesicles, their electrostatic attraction at the vesicle interface, possible cross-linking with other vesicles and different hydration features of the isomeric precursors of the C-dots. The study may be helpful in understanding the interactions and attachment processes of C-dots at the interface of biological membranes.

Interaction of aliphatic amino acids with zwitterionic and charged lipid membranes: hydration and dehydration phenomena.

In the present contribution, we investigate the interactions of lipid bilayer membranes of different charges and different phase states with aliphatic amino acids of varying charge (aspartic acid, glutamic acid, arginine and lysine) and hydrophobicity (serine, leucine and valine) by steady state and time-resolved spectroscopic techniques, dynamic light scattering (DLS) measurements and confocal imaging (CLSM). The study reveals that negatively charged amino acids such as aspartic acid and glutamic acid interact strongly with the lipid membranes particularly with negatively charged lipid membranes by stabilizing their gel phase. On the other hand, positively charged amino acids bring in hydration in the membranes. We explain this unique observation by the shift in pKa of amino acids in the vicinity of the lipid membranes and solvation and desolvation processes in the light of recent computer simulations. We also find that hydrogen bonding plays a significant role in governing the interaction of aliphatic amino acids with zwitterionic lipid membranes. The more polar serine bearing a hydroxyl group at the terminal carbon offers a stronger interaction with the lipid bilayer membranes as compared to its analogues leucine and valine, which are hydrophobic in nature.

Influence of Trivalent Metal Ions on Lipid Vesicles: Gelation and Fusion Phenomena.

In this contribution, we report the interaction of 1,2-dimyristoyl- sn-glycero-3-phosphocholine (DMPC) lipid vesicles with a series of trivalent metal ions of the same group, namely, Al3+, Ga3+, and In3+, to get a distinct view of the effect of size, effective charge, and hydration free energy of these metal ions on lipid vesicles. We employed steady-state and time-resolved spectroscopic techniques including time-resolved anisotropy measurement, confocal imaging, and dynamic light scattering (DLS) measurement to probe the interaction. Our study reveals that all of the three trivalent metal ions induce gelation in lipid vesicles by removing water molecules from the interfacial region. The extent of gelation induced by the metal ions follows the order of In3+ > Ga3+ ≥ Al3+. We explain this observation in light of different free-energy terms. Notably, the degree of interaction for trivalent metal ions is higher as compared to that for divalent metal ions at physiological pH (pH ∼ 7.0). Most importantly, we observe that unlike divalent metal ions, trivalent metal ions dehydrate the lipid vesicles even at lower pH. The DLS measurement and confocal imaging indicate that In3+ causes significant aggregation or fusion of the PC vesicles, while Al3+ and Ga3+ did not induce any aggregation at the experimental concentration. We employ Derjaguin-Landau-Vervey-Overbeek (DLVO) theory to explain the aggregation phenomena induced by In3+.

Effect of Surface Ligand and Temperature on Lipid Vesicle-Gold Nanoparticle Interaction: A Spectroscopic Investigation.

We herein investigate the interactions of differently functionalized anionic and cationic gold nanoparticles (AuNPs) with zwitterionic phosphocholine (PC) as well as inverse phosphocholine (iPC) lipid bilayers via spectroscopic measures. In this study, we used PC lipids with varying phase-transition temperatures, i.e., DMPC ( Tm = 24 °C), DOPC ( Tm = -20 °C), and iPC lipid DOCP ( Tm = -20 °C) to study their interactions with AuNPs functionalized with anionic ligands citrate, 3-mercaptopropionic acid, glutathione, and cationic ligand cysteamine. We studied the interactions by steady-state and time-resolved spectroscopic studies using membrane-sensitive probes 6-propionyl-2-dimethylaminonaphthalene (PRODAN) and 8-anilino-1 naphthalenesulfonate (ANS), as well as by confocal laser scanning microscopy (CLSM) imaging and dynamic light scattering (DLS) measurements. We observe that AuNPs bring in stability to the lipid vesicle, and the extent of interaction differs with the different surface ligands on the AuNPs. We observe that AuNPs functionalized with citrate effectively increase the phase-transition temperature of the vesicles by interacting with them. Our study reveals that the extent of interaction depends on the bulkiness of the ligands attached to the AuNPs. The bulkier ligands exert less van der Waals force, resulting in a weaker interaction. Moreover, we find that the interactions are more strongly pronounced when the vesicles are near the phase-transition temperature of the lipid.  The CLSM imaging and DLS measurements demonstrate the surface modifications in the vesicles as a result of these interactions.

Spectroscopic evidence for hydration and dehydration of lipid bilayers upon interaction with metal ions: a new physical insight.

In this manuscript, we investigate the interactions of different metal ions with zwitterionic phospholipid bilayers of different chain lengths using the well-known membrane probe PRODAN and steady state and time resolved fluorescence spectroscopy. We used three zwitterionic lipids that are widely different in their phase transition temperature, namely, dipalmitoylphosphatidylcholine (DPPC), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), and 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine (POPC) and salts of zinc (Zn), calcium (Ca) and magnesium (Mg). The steady state and time resolved studies reveal that the affinity of the metal ions follows the order Zn2+ > Ca2+ > Mg2+. The study further reveals that the lipid membrane with an unsaturated chain exhibits very small affinity towards metal ions. We find that the Zn2+ and Ca2+ metal ions induce significant gelation in the lipid bilayer possibly by dehydrating the lipid bilayer surface. The study also demonstrates that unlike Zn2+ and Ca2+, dehydration does not take place for Mg2+. The extreme hydration induced by Mg2+ is rationalized by the tight hydration of Mg2+ and very high free energy barrier of Mg2+ to bind with lipid oxygen as compared to that of water molecules.

Spectroscopic Study of the Interaction of Carboxyl-Modified Gold Nanoparticles with Liposomes of Different Chain Lengths and Controlled Drug Release by Layer-by-Layer Technology.

In this article, we investigate the interactions of carboxyl-modified gold nanoparticles (AuC) with zwitterionic phospholipid liposomes of different chain lengths using a well-known membrane probe PRODAN by steady-state and time-resolved spectroscopy. We use three zwitterionic lipids, namely, dipalmitoylphosphatidylcholine (DPPC), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), and 1,2-dilauroyl-sn-glycero-3-phosphocholine (DLPC), which are widely different in their phase transition temperatures to form liposome-AuC assemblies. The steady-state and time-resolved studies indicate that the AuC brings in stability toward liposomes by local gelation. We observe that the bound AuC detach from the surface of the liposomes under pH ≈ 5 due to protonation of the carboxyl group, thus eliminating the electrostatic interaction between nanoparticles and head groups of liposomes. The detachment rate of AuC from the liposome-AuC assemblies is different for the aforementioned liposomes due to differences in their fluidity. We exploited the phenomena for the controlled release of a prominent anticancer drug Doxorubicin (DOX) under acidic conditions for different zwitterionic liposomes. The drug release rate was further optimized by coating of liposome-AuC assemblies with oppositely charged polymer (P), polydiallyldimethylammonium chloride, followed by a mixture of lipids L (DMPC:DMPG) and again with a polymer in a layer-by-layer fashion to obtain capsule-like structures. This system is highly stable for weeks, as confirmed by field-emission scanning electron microscopy (FE-SEM) and confocal laser scanning microscopy (CLSM) imaging, and inhibits premature release. The layer coating was confirmed by hydrodynamic size and zeta potential measurements of the systems. The capsules obtained are of immense importance as they can control release of the drug from the systems to a large extent.