Protein vaporization

Protein Vapor diffusion hanging crystallization drops Chayen etal. (2001)... 

Many complex systems have been spread on liquid interfaces for a variety of reasons. We begin this chapter with a discussion of the behavior of synthetic polymers at the liquid-air interface. Most of these systems are linear macromolecules however, rigid-rod polymers and more complex structures are of interest for potential optoelectronic applications. Biological macromolecules are spread at the liquid-vapor interface to fabricate sensors and other biomedical devices. In addition, the study of proteins at the air-water interface yields important information on enzymatic recognition, and membrane protein behavior. We touch on other biological systems, namely, phospholipids and cholesterol monolayers. These systems are so widely and routinely studied these days that they were also mentioned in some detail in Chapter IV. The closely related matter of bilayers and vesicles is also briefly addressed. 

The ablated vapors constitute an aerosol that can be examined using a secondary ionization source. Thus, passing the aerosol into a plasma torch provides an excellent means of ionization, and by such methods isotope patterns or ratios are readily measurable from otherwise intractable materials such as bone or ceramics. If the sample examined is dissolved as a solid solution in a matrix, the rapid expansion of the matrix, often an organic acid, covolatilizes the entrained sample. Proton transfer from the matrix occurs to give protonated molecular ions of the sample. Normally thermally unstable, polar biomolecules such as proteins give good yields of protonated ions. This is the basis of matrix-assisted laser desorption ionization (MALDI). 

Films or membranes of silkworm silk have been produced by air-drying aqueous solutions prepared from the concentrated salts, followed by dialysis (11,28). The films, which are water soluble, generally contain silk in the silk I conformation with a significant content of random coil. Many different treatments have been used to modify these films to decrease their water solubiUty by converting silk I to silk II in a process found usehil for enzyme entrapment (28). Silk membranes have also been cast from fibroin solutions and characterized for permeation properties. Oxygen and water vapor transmission rates were dependent on the exposure conditions to methanol to faciUtate the conversion to silk II (29). Thin monolayer films have been formed from solubilized silkworm silk using Langmuir techniques to faciUtate stmctural characterization of the protein (30). ResolubiLized silkworm cocoon silk has been spun into fibers (31), as have recombinant silkworm silks (32). 

Figure 18.4 The hanging-drop method of protein crystallization, (a) About 10 pi of a 10 mg/ml protein solution in a buffer with added precipitant—such as ammonium sulfate, at a concentration below that at which it causes the protein to precipitate—is put on a thin glass plate that is sealed upside down on the top of a small container. In the container there is about 1 ml of concentrated precipitant solution. Equilibrium between the drop and the container is slowly reached through vapor diffusion, the precipitant concentration in the drop is increased by loss of water to the reservoir, and once the saturation point is reached the protein slowly comes out of solution. If other conditions such as pH and temperature are right, protein crystals will occur in the drop, (b) Crystals of recombinant enzyme RuBisCo from Anacystis nidulans formed by the hanging-drop method. (Courtesy of Janet Newman, Uppsala, who produced these crystals.)...

It follows, that the peak width of a solute could give an indication of its molecular weight and, although the data may not be precise, approximate values could be extremely valuable when dealing with very large molecular weight substances such as polypeptides and proteins. In particular, the technique would be very useful for those substances that are extremely difficult, or impossible, to vaporize in the ion source of a mass spectrometer to provide mass data. 

MALDI is a LIMS method capable of vaporizing and ionizing large biological molecules such as proteins or DNA. The biological molecules are dispersed in a solid matrix that serves as a carrier. 

Cormier and Dure (1963) found another type of luciferin and called it protein-free luciferin. Protein-free luciferin was found in the vapor condensate of freeze-drying whole animals, and also in the 3 5-56 % ammonium sulfate fraction of the crude extract noted above. The protein-free luciferin behaved like an aromatic or heterocyclic compound and it was strongly adsorbed onto Sephadex and other chromatography media, requiring a considerable amount of solvent to elute it. The luminescence reaction of protein-free luciferin in the presence of luciferase required a 500-times higher concentration of H2O2 compared with the standard luciferin preparation. Both types of the luciferin preparation had a strong odor of iodoform. 

In addition to chemical reactions, the isokinetic relationship can be applied to various physical processes accompanied by enthalpy change. Correlations of this kind were found between enthalpies and entropies of solution (20, 83-92), vaporization (86, 91), sublimation (93, 94), desorption (95), and diffusion (96, 97) and between the two parameters characterizing the temperature dependence of thermochromic transitions (98). A kind of isokinetic relationship was claimed even for enthalpy and entropy of pure substances when relative values referred to those at 298° K are used (99). Enthalpies and entropies of intermolecular interaction were correlated for solutions, pure liquids, and crystals (6). Quite generally, for any temperature-dependent physical quantity, the activation parameters can be computed in a formal way, and correlations between them have been observed for dielectric absorption (100) and resistance of semiconductors (101-105) or fluidity (40, 106). On the other hand, the isokinetic relationship seems to hold in reactions of widely different kinds, starting from elementary processes in the gas phase (107) and including recombination reactions in the solid phase (108), polymerization reactions (109), and inorganic complex formation (110-112), up to such biochemical reactions as denaturation of proteins (113) and even such biological processes as hemolysis of erythrocytes (114). 

Biodegradable films made from edible biopolymers from renewable sources could become an important factor in reducing the environmental impact of plastic waste. Proteins, lipids, and polysaccharides are the main biopolymers employed to make edible films and coatings. Which of these components are present in different proportions and determine the properties of the material, as a barrier to water vapor, oxygen, carbon dioxide, and lipid transfer in food systems (Gomez-Guillen et al. 2002 and 2009). 

The amount of water boimd to the proteins and polysaccharides depends primarily on the ratio of water to the biopolymer in the investigated system The two extreme cases are the dry biopolymer (water content tend to zero) and highly diluted aqueous solutions of the biopolymers. The dry biopolymer undergoes hydration if is exposed to the water vapor of increased vapor pressure. The extent of hydration can be determined y measuring the... 

D. E. Overcashier, T. W. Patapoff, and C. C. Hsu, Lyophilization of protein formulations in vials Investigation of the relationship between resistance to vapor flow during primary drying and small scale product collapse, J. Pharm. Sci., 88(7), 688 (1999). 

Figure 3 Protein crystallization diagrammatic representation of the hanging-drop method of vapor diffusion.

Purified MeHNL was crystallized by the sitting-drop vapor-diffusion method. The 10-20 mm bipyramidal crystals formed were cross-linked with glutaraldehyde and used as biocatalyst for the synthesis of optically active cyanohydrins. The cross-linked crystals were more stable than Celite-immobilized enzymes when incubated in organic solvents, especially in polar solvents. After six consecutive batch reactions in dibutyl ether, the remaining activity of the cross-linked crystals was more than 70 times higher than for the immobilized enzymes. Nevertheless, the specific activity of the cross-linked crystals per milligram protein was reduced compared with the activity of Celite-immobilized enzymes [53],... 

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