PH-gradient formation

During the course of the pH-gradient formation, the conductance will decrease and vary from anode to cathode. This is due to the following facts ... 

Capillary isoelectric focusing (CIEF) is designed to separate solutes based on their p/, the pH where they are electrically neutral. Because solutes do not migrate when they are neutral, the mobilization step distinguishes CIEF from other capillary electrophoretic techniques. This entry reviews the basis for CIEF including pH gradient formation, mobilization techniques, additives, and applications. 

Figure 3. Kinetics for the pH Gradient Formation Due to a KCl Concentration Difference. The filter was impregnated with oleic acid (Adapted from ref 4.).

ELECTRIC POTENTIAL AND pH GRADIENT FORMATION BY RECONSTITUTED ATPase PROTEO-LIPOSOMES FROM THE THERMOPHILIC CYANOBACTERIUM SYNECHOCOCCUS 6716... 

Diluted in neutraUzed water, separation on CIS Novapak using gradient elution using methanol and phosphate buffers at pH 7 Formation of ion pairs with cetylpyridinium chloride (CeCl) in water and extraction in butanol, separation on C8 Spherisorb using gradient of acetonitrile, methanol, CeCl/phosphate buffer... 

BLM transport systems for ferrioxamine B were also devised based on first coordination shell recognition via ternary complex formation utilizing vacant coordination sites on the Fe(III) center (Fig. 29) (199). The tetra-coordinated substrate complex selectively transported was partially dechelated diaqua-ferrioxamine B and coordinately unsaturated di-hydroxamato iron(III) complexes, which utilized a hydrophobic membrane bound bidentate chelator as a carrier for selective transport. Active transport for these systems was accomplished using a pH gradient (199). 

IEF is similar in concept to conventional gel IEF a stable pH gradient is formed in the capillary using carrier ampholytes, and proteins are focused in the gradient at their pis. The major difference in performing IEF in the capillary format rather than slab gel is the requirement for mobilizing focused protein zones past the detection point. IEF is described in Capillary Isoelectronic Focusing. ... 

If the second dialysis step against external buffer is omitted during the formation of LUV, transmembrane pH gradients can be formed by running... 

Further work found that the formation of acidic microdomains occurs at the stratum granulosum-SC interface and is regulated by the sodium-proton exchanger NHEl. In addition, the acidic SC surface is not fully developed at birth, and rather acidic microdomains at the SC-SG interface develop postnatally (Behne et al. 2003). Niesner et al. also used TP FLIM on artificial skin constructs and found that an identical pH gradient to that found in mammalian skin exists, which could enable further research on barrier function without the need for human or animal tissues (Niesner et al. 2005). 

The peptide subunit was easily incorporated into lipid bilayers of liposome, as confirmed by absorption and fluorescence spectroscopy. Formation of H-bonded transmembrane channel structure was confirmed by FT IR measurement, which suggests the formation of a tight H-bond network in phosphatidylcholine liposomes. Liposomes were first prepared to make the inside pH 6.5 and the outside pH 5.5. Then the addition of the peptide to such liposomal suspensions caused a rapid collapse of the pH gradient. The proton transport activity was comparable to that of antibiotics gramicidin A and amphotericin B. 

A crucially important finding is that submitochon-drial particles or vesicles from broken chloroplasts will synthesize ATP from ADP and P , when an artificial pH gradient is imposed.172186 Isolated purified FjF0 ATPase from a thermophilic Bacillus has been coreconstituted into liposomes with the light-driven proton pump bacteiiorhodopsin (Chapter 23). Illumination induced ATP synthesis.187 These observations support Mitchell s proposal that the ATP synthase is both spatially separate from the electron carriers in the membrane and utilizes the protonmotive force to make ATP. Thus, the passage of protons from the outside of the mitochondria back in through the ATP synthase induces the formation of ATP. What is the stoichiometry of this process ... 

The chemiosmotic theory postulates that protons moving back into the matrix via an ATP-synthase drive the formation of ATP. Evidence for this is that an electrochemical potential gradient for protons can support the formation of ATP in the absence of electron-transfer reactions. A transient pH gradient that pulls protons into the matrix can be set up by first incubating mitochondria at pH 9, so that the inside becomes alkaline, and then quickly lowering the pH of the suspension medium to 7 (fig. 14.21). 

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