Transitions protein phase

Native biological membranes also display characteristic phase transitions, but these are broad and strongly dependent on the lipid and protein composition of the membrane. 

Bilayer phase transitions are sensitive to the presence of solutes that interact with lipids, including multivalent cations, lipid-soluble agents, peptides, and proteins. 

Discuss the effects on the lipid phase transition of pure dimyris-toyl phosphatidylcholine vesicles of added (a) divalent cations, (b) cholesterol, (c) distearoyl phosphatidylserine, (d) dioleoyl phosphatidylcholine, and (e) integral membrane proteins. 

Membranes are composed of phospholipids and proteins. The fatty acid composition of the phospholipids in a membrane influences how it is affected by the cold. In general, as the temperature of a cell is lowered the lipids in the membrane bilayer undergo a phase transition from a liquid crystalline (fluid) state to a gel (more solid) state. The temperature at which this transition takes place is very narrow for phospholipids composed of a simple mixture of fatty acids, but is quite broad for the phospholipids in cellular membranes. It is usually implied from various methods... 

Hypothermia—Indirect cryodestruction Metabolic uncoupling Energy deprivation Ionic imbalance Disruption of acid-base balance Waste accumulation Membrane phase transitions Cytoskeletal disassembly Frozen State—Direct cryodestruction Water solidification Hyperosmolality Cell-volume disruption Protein denaturation Tissue shearing Intracellular-ice propagation Membrane disruption Microvascular Thawed State Direct effects... 

Applying MD to systems of biochemical interest, such as proteins or DNA in solution, one has to deal with several thousands of atoms. Models for systems with long spatial correlations, such as liquid crystals, micelles, or any system near a phase transition or critical point, also must involve a large number of atoms. Some of these systems, including synthetic polymers, obey certain scaling laws that allow the estimation of the behaviour of a large system by extrapolation. Unfortunately, proteins are very precise structures that evade such simplifications. So let us take 10,000 atoms as a reasonable size for a realistic complex system. 

Fig. 9 Purification of ELPs by ITC is based on the reversible inverse phase transition. Le/i Protein purification via direct ELP fusions. A soluble ELP fused to a target protein becomes reversibly insoluble upon increasing temperature above 7,. Center Protein purification via ELP coaggregation. An excess of free ELPs enhances the aggregation of trace quantities of ELP-fusions. Right Purification via ELP-mediated affinity capture (EMAC). ELPs are fused to capture proteins, which bind specifically and reversibly to a target protein. This target protein can then be aggregated at temperatures above the T,. Adapted from [38] with permission from Elsevier, copyright 2010...

In a subsequent study, the effect of reducing the ELP molecular weight on the expression and purification of a fusion protein was investigated. Two ELPs, ELP [V-20] and ELP[VsA2G3-90], both with a transition temperature at 40°C in phosphate-buffered saline (PBS) containing 1 M NaCl, were applied for the purification of thioredoxin. Similar yields were observed for both fusion proteins, resulting in a higher thioredoxin yield for the ELP[V-20] fusion, since the ELP fraction was smaller. However, a more complex phase transition behavior was observed for this ELP and therefore a selection of an appropriate combination of salt concentration and solution temperature was required [39]. 

The previous ELP fusions all are examples of protein purification in which the ELP is covalently connected to the protein of choice. This approach is suitable for the purification of recombinant proteins that are expressed to high levels, but at very low concentrations of ELP the recovery becomes limited. Therefore this approach is not applicable for proteins expressed at micrograms per liter of bacterial culture, such as toxic proteins and complex multidomain proteins. An adjusted variant of ITC was designed to solve this problem. This variant makes use of coaggregation of free ELPs with ELP fusion proteins. In this coaggregation process, an excess of free ELP is added to a cell lysate to induce the phase transition at low concentrations of... 

Conformational and phase transitions can potentially be indicative of the primary structure of thermosensitive macromolecules. Indeed, depending on the relative location of H- and P-blocks, as well as on the variation of their length, the chains can either undergo conformational transition accompanied by phase separation, or they can exhibit only the conformational changes without macroscopic phase transitions, i.e. the behaviour observed in the case of protein-like HP-copolymers. Therefore, the solution behaviour of separated fractions of these NVCl/NVIAz-copolymers in an aqueous medium at different temperatures is very important. 

Polymers designed with this technique have a number of important aspects in common with proteins. First of all, the transition from a liquid-like globule into a frozen state occurs as a first order phase transition. Further, the frozen state itself has an essential stability margin, which is determined by the design parameters. As in real proteins, neither a large variation of temperature or other environmental conditions, nor a mutational substitution of several monomers leads to any change in basic state conformation. In this respect the ability of sequence design to capture certain essential characteristics of proteins seems quite plausible. 

Murray and Hunt, 1993). Cyclins, kinases, and phosphatases that regulate the passage of the cell through the G] — S phase transition are all present in mammals, invertebrates, and plants (Solomon, 1993 Doonan and Fobart, 1997 Zavitz and Zipursky, 1997). However, multicellular eukaryotes contain multiple orthologs of yeast cell cycle proteins they initiate proliferation via growth factors, rather than, for example, yeast mating factors, and they possess additional checkpoint controls and repair pathways. 

The fluorescence lifetime is sensitive to the environment of the fluorophore, and in membranes this usually means the surrounding fatty acyl chains or the membrane protein interfacial region (see summary in Table 5.3). Generally, the lifetime of membrane-bound fluorophores is rather less sensitive to the types of subtle alterations which are encountered in membranes as compared to the fluorescence anisotropy parameters. The gel-to-liquid crystalline phase transition is a notable exception where most fluorophores show an alteration in lifetime properties. Although, again, the anisotropy (see below) is the most sensitive parameter in this regard, the fluorescence lifetime has been used with considerable success in the study of phase transitions and lateral phase separations. Fluorophores used to yield information on the... 

Papahajopoulos, D., M. Moscarello, E.H. Eylar, and T. Isac. 1975. Effects of proteins on thermotropic phase transitions of phospholipid membranes. Biochim Biophys Acta 401 317-335. 

Mathias, N., et al., Cdc53p acts in concert with Cdc4p and Cdc34p to control the Gl-to-S-phase transition and identifies a conserved family of proteins. Mol Cell Biol, 1996, 16(12), 6634-43. 

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