scholarly journals Xanthene-dye-labelled phosphatidylethanolamines as probes of interfacial pH. Studies in phospholipid vesicles

1989 ◽  
Vol 258 (3) ◽  
pp. 683-687 ◽  
Author(s):  
C G Knight ◽  
T Stephens
Keyword(s):  
Phospholipid Vesicles ◽  
Amino Group ◽  
Cell Surfaces ◽  
Vesicle Membrane ◽  
Xanthene Dyes ◽  
Ph Values ◽  
Ph Indicators ◽  
Membrane Bound ◽  
Single Emission ◽  
Hasselbalch Equation

We have been developing the use of plasma-membrane-bound fluorescent probes to measure the pH values at the surfaces of living chondrocytes. For this purpose, three lipophilic pH indicators were made by covalently binding the xanthene dyes fluorescein, eosin or dichlorofluorescein to the amino group of phosphatidylethanolamine. The probes were incorporated into phospholipid vesicles and the effect of pH on the fluorescence was characterized. Fluorescence was measured at a single emission wavelength during excitation at two wavelengths, and the ratio of the intensities was calculated. The experimentally observed pKobs. values were determined by fitting the fluorescence ratios to the Henderson-Hasselbalch equation. All three probes acted as pH indicators, and the eosinyl-, dichlorofluoresceinyl- and fluoresceinylphosphatidylethanolamines had pKobs. values of 3.5, 6.3 and 7.5 respectively. At physiological salt concentrations, changes in the composition of the vesicle membrane had little effect on these values. We concluded that these probes were promising candidates for the measurement of pH values at cell surfaces.

scholarly journals Selective extraction of membrane-bound proteins by phospholipid vesicles.

1977 ◽  
Vol 252 (19) ◽  
pp. 6759-6763 ◽  
Author(s):  
S R Bouma ◽  
F W Drislane ◽  
W H Huestis
Keyword(s):  
Selective Extraction ◽  
Phospholipid Vesicles ◽  
Membrane Bound

Reaction of metallic nitrilotriacetates with histamine

1969 ◽  
Vol 47 (8) ◽  
pp. 1269-1273 ◽  
Author(s):  
A. L. Beauchamp ◽  
J. Israeli ◽  
H. Saulnier
Keyword(s):  
Potentiometric Titration ◽  
Formation Constants ◽  
Amino Group ◽  
Mixed Complex ◽  
Ph Values ◽  
Titration Curves ◽  
Terminal Amino ◽  
Complex Ion

Cu(II), Ni(II), Co(II), and Zn(II) nitrilotriacetates (MeX−) react with histamine nitrate (LH+) to form a protonated mixed complex MeXLH where the metal appears to be bound only to the tertiary imidazolic nitrogen of histaminium ion. At higher pH values the proton dissociates to yield a mixed complex ion MeXL− in which both the imidazolic nitrogen and the terminal amino group are coordinated. The formation constants of these species were calculated from the potentiometric titration curves.

scholarly journals Fusion of phospholipid vesicles with a planar membrane depends on the membrane permeability of the solute used to create the osmotic pressure.

1989 ◽  
Vol 93 (2) ◽  
pp. 201-210 ◽  
Author(s):  
F S Cohen ◽  
W D Niles ◽  
M H Akabas
Keyword(s):  
Osmotic Pressure ◽  
Lipid Bilayer ◽  
Lipid Membrane ◽  
Contact Region ◽  
Companion Paper ◽  
Phospholipid Vesicles ◽  
Osmotic Gradient ◽  
Vesicle Membrane ◽  
Permeable Channel ◽  
Planar Membrane

Phospholipid vesicles fuse with a planar membrane when they are osmotically swollen. Channels in the vesicle membrane are required for swelling to occur when the vesicle-containing compartment is made hyperosmotic by adding a solute (termed an osmoticant). We have studied fusion using two different channels, porin, a highly permeable channel, and nystatin, a much less permeable channel. We report that an osmoticant's ability to support fusion (defined as the magnitude of osmotic gradient necessary to obtain sustained fusion) depends on both its permeability through lipid bilayer as well as its permeability through the channel by which it enters the vesicle interior. With porin as the channel, formamide requires an osmotic gradient about ten times that required with urea, which is approximately 1/40th as permeant as formamide through bare lipid membrane. When nystatin is the channel, however, fusion rates sustained by osmotic gradients of formamide are within a factor of two of those obtained with urea. Vesicles containing a porin-impermeant solute can be induced to swell and fuse with a planar membrane when the impermeant bathing the vesicles is replaced by an isosmotic quantity of a porin-permeant solute. With this method of swelling, formamide is as effective as urea in obtaining fusion. In addition, we report that binding of vesicles to the planar membrane does not make the contact region more permeable to the osmoticant than is bare lipid bilayer. In the companion paper, we quantitatively account for the observation that the ability of a solute to promote fusion depends on its permeability properties and the method of swelling. We show that the intravesicular pressure developed drives fusion.

Photometric Determination of pH with a Single Standard and Calculation by Nomogram

1966 ◽  
Vol 12 (12) ◽  
pp. 849-870 ◽  
Author(s):  
Donald D Van Slyke ◽  
Lawrence V Hankes ◽  
Janis John Vitols
Keyword(s):  
Human Plasma ◽  
Photometric Determination ◽  
Glass Electrode ◽  
Phenol Red ◽  
Ph Values ◽  
Slow Fading ◽  
Standard Solution ◽  
Single Standard ◽  
Hasselbalch Equation

Abstract A method is described for construction of a nomogram, based on the Henderson Hasselbalch equation, with which photometric pH values can be calculated from the absorbance of an indicator in a sample and the absorbance of the indicator in a single standard solution. Thereby the necessity of preparing calibration curves from a series of standard solutions is avoided. The procedure is particularly convenient when the stock solution of the indicator is subject to slow fading, as in the case of phenol red. An application of the procedure to the photometric determination of the pH of human plasma is detailed and the results are compared with those obtained with a glass electrode.

scholarly journals Modification of ryanodine receptor/Ca2+ release channel with dinitrofluorobenzene

1999 ◽  
Vol 342 (1) ◽  
pp. 239-248 ◽  
Author(s):  
Nurit HADAD ◽  
Wei FENG ◽  
Varda SHOSHAN-BARMATZ
Keyword(s):  
Ryanodine Receptor ◽  
Single Channel ◽  
Ruthenium Red ◽  
Amino Group ◽  
Channel Activity ◽  
Closed Channel ◽  
Release Channel ◽  
Atp Binding Site ◽  
Ph Values ◽  
Stimulation Of

Modification of the ryanodine receptor (RyR)/Ca2+ release channel with 2,4-dinitrofluorobenzene (DNFB) indicated that two classes of amino group interact with the reagent, as can be distinguished on the basis of their reactivity/accessibility and the effects on ryanodine binding and single channel activities. One group interacted very rapidly (t½ < 30 s) at 25 °C with low concentrations of DNFB [C50 (concentration of DNFB required for 50% inhibition or stimulation of ryanodine binding) = 5 μM], and at pH values of 6.2 and higher. This interaction resulted in the marked stimulation of ryanodine binding and the complete inhibition of a single Ca2+ release channel incorporated into planar lipid bilayer. The second group is accessible at higher temperatures (37 °C); at pH values higher than 7.4 it reacted slowly (t½ = 20 min) with high concentrations of DNFB (C50 = 70 μM). This interaction led to the inhibition of ryanodine binding and single channel activity. Modification of RyR with DNFB under the stimulatory conditions resulted in 3.6-fold and 6-fold increases in ryanodine-binding and Ca2+-binding affinities respectively. Modification with DNFB under the inhibitory conditions resulted in a decrease in the total ryanodine-binding sites. The exposure of the RyR single channel to DNFB under both inhibitory and stimulatory conditions led to the complete closure of the channel. However, when modified under the stimulatory conditions, but not under the inhibitory ones, the DNFB-modified closed channel could be re-activated by sub-micromolar concentrations of ryanodine, in the presence of nanomolar concentrations of Ca2+. The DNFB-modified ryanodine-activated RyR channel showed fast transitions between open, closed and several sub-conductance states, and was completely closed by Ruthenium Red. ATP re-activated the DNFB-modified closed channel or, if present during modification, prevented the inhibition of RyR channel activity by DNFB. Neither the stimulation nor the inhibition of ryanodine binding by modification with DNFB was affected by the presence of ATP. By using the photoreactive ATP analogue 3′-O-(4-benzoyl)benzoyl-[α-32P]ATP we found that DNFB modification had no effect on the ATP-binding site of RyR. The results are discussed with regard to the involvement of amino group residues in channel gating, ryanodine association/dissociation and occlusion, and the relationship between the open/closed state of the RyR and its capacity to bind ryanodine.

Roles of lysine residues and N-terminal α-amino group in membrane-damaging activity of Taiwan cobra cardiotoxin 3 toward anionic and zwitterionic phospholipid vesicles

Toxicon ◽  
2010 ◽  
Vol 55 (2-3) ◽  
pp. 256-264 ◽  
Author(s):  
Yi-Ling Chiou ◽  
Pei-Hsiu Kao ◽  
Wen-Hsin Liu ◽  
Shinne-Ren Lin ◽  
Long-Sen Chang
Keyword(s):  
Phospholipid Vesicles ◽  
Amino Group ◽  
Lysine Residues

Heterogeneity of Surfaces of Subgingival Bacteria as Detected by Zeta Potential Measurements

1992 ◽  
Vol 71 (11) ◽  
pp. 1803-1806 ◽  
Author(s):  
M.M. Cowan ◽  
H.C. Van der Mei ◽  
I. Stokroos ◽  
H.J. Busscher
Keyword(s):  
Ph Dependence ◽  
Laboratory Strain ◽  
Prevotella Intermedia ◽  
Gram Negative Bacteria ◽  
Cell Surfaces ◽  
Fluid Medium ◽  
Gram Negative ◽  
Zeta Potentials ◽  
Ph Values ◽  
Ph Dependent

Porphyromonasgingivalis, Prevotella intermedia, and Actinobacillus actinomycetemcomitans (A.a.) are Gram-negative bacteria which are implicated in various forms of periodontal disease. The Gram-positive Peptostreptococcus micros may also play an important role. For investigation of the possible adhesion and colonization mechanisms of these organisms, the charge properties of the outermost layers of bacterial cell surfaces were studied through the measurement of zeta potentials at various pH values. Eleven fresh clinical isolates, representing the four species, and one laboratory strain, P. gingivalis W83, were examined. Eleven of the 12 strains displayed heterogeneity with respect to pH-dependent zeta potentials. Within single cultures of each of these strains, two distinct populations of cells were found, one which was more negatively charged than the other. For the Gram-negative strains, the more negatively charged subpopulation was in the majority, while the P. micros strains appeared to be composed mainly of a less-negatively-charged subpopulation. Vesicles prepared from two strains displayed the same pH dependence and heterogeneity of zeta potentials as the parent cells. An A.a. strain which was passaged several times in fluid medium had lost its fimbriae and became homogeneous with respect to charge.

Kinetics of reaction of 1,2-diaminobenzene with phenylglyoxal

1990 ◽  
Vol 55 (5) ◽  
pp. 1216-1222 ◽  
Author(s):  
Jiří Klicnar ◽  
Jaromír Mindl ◽  
Vojeslav Štěrba
Keyword(s):  
Reaction Kinetics ◽  
Cyclization Reaction ◽  
Amino Group ◽  
Ph Values ◽  
Rate Limiting Step ◽  
Limiting Step ◽  
Kinetics Of Reaction ◽  
Rate Limiting ◽  
Kinetics Of ◽  
Aldehydic Group

The cyclization reaction kinetics of phenylglyoxal monohydrate with 1,2-diaminobenzene have been studied in formate, acetate, and phosphate buffers. At high pH values and low buffer concentrations the rate-limiting step consists in the protonation of the intermediate formed by addition of the first amino group to aldehydic group of phenylglyoxal. With increasing concentrations of formate and acetate buffers the rate-limiting step shifts to the formation of the intermediate. In phosphate buffers the catalysis by the basic buffer component makes itself felt, too. At higher concentrations of 1,2-diaminobenzene, the dehydration of phenylglyoxal monohydrate gradually becomes the rate-limiting step.

Diphosphoinositide metabolism in bovine adrenal medulla

1976 ◽  
Vol 54 (8) ◽  
pp. 746-753 ◽  
Author(s):  
Y. A. Lefebvre ◽  
D. A. White ◽  
J. N. Hawthorne
Keyword(s):  
Adrenal Medulla ◽  
Calcium Binding ◽  
Microsomal Fraction ◽  
Kinase Activity ◽  
Hydrolytic Activity ◽  
Vesicle Membrane ◽  
Bovine Adrenal Medulla ◽  
Phosphatidylinositol Kinase ◽  
Membrane Bound ◽  
Phosphatidylinositol 4

(1) A phosphatidylinositol kinase (EC 2.7.1.67) ofa chromaffin vesicle membrane preparation isolated from bovine adrenal medulla was characterized. Its activity towards endogenous and exogenous phosphatidylinositol was very similar to the kinase activity of the microsomal fraction prepared from the same tissue.(2) Phosphomonoesterase (EC 3.1.3.36) and diesterase activity hydrolysing membrane bound phosphatidylinositol 4-phosphate was located mainly in the microsomal fraction. No hydrolytic activity was present in the vesicle membrane.(3) Phosphorylation of chromaffin vesicle membrane phosphatidylinositol did not increase calcium-binding by the membranes.

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