steady-state pH gradient Dotaz Zobrazit nápovědu
In 1961, Svensson described isoelectric focusing (IEF), the separation of ampholytic compounds in a stationary, natural pH gradient that was formed by passing current through a sucrose density gradient-stabilized ampholyte mixture in a constant cross-section apparatus, free of mixing. Stable pH gradients were formed as the electrophoretic transport built up a series of isoelectric ampholyte zones-the concentration of which decreased with their distance from the electrodes-and a diffusive flux which balanced the generating electrophoretic flux. When polyacrylamide gel replaced the sucrose density gradient as the stabilizing medium, the spatial and temporal stability of Svensson's pH gradient became lost, igniting a search for the explanation and mitigation of the loss. Over time, through a series of insightful suggestions, the currently held notion emerged that in the modern IEF experiment-where the carrier ampholyte (CA) mixture is placed between the anolyte- and catholyte-containing large-volume electrode vessels (open-system IEF)-a two-stage process operates that comprises a rapid first phase during which a linear pH gradient develops, and a subsequent slow, second stage, during which the pH gradient decays as isotachophoretic processes move the extreme pI CAs into the electrode vessels. Here we trace the development of the two-stage IEF model using quotes from the original publications and point out critical results that the IEF community should have embraced but missed. This manuscript sets the foundation for the companion papers, Parts 2 and 3, in which an alternative model, transient bidirectional isotachophoresis is presented to describe the open-system IEF experiment.
A model was constructed which includes electron transport (linear and cyclic and Mehler type reaction) coupled to proton translocation, counter ion movement, ATP synthesis, and Calvin-Benson cycle. The focus is on modeling of the light-induced total electric potential difference (ΔΨ) which in this model originates from the bulk phase electric potential difference (ΔΨb), the localized electric potential difference (ΔΨc), as well as the surface electric potential difference (ΔΨs). The measured dual wavelength transmittance signal (ΔA515-560nm, electrochromic shift) was used as a proxy for experimental ΔΨ. The predictions for theoretical ΔΨ vary with assumed contribution of ΔΨs, which might imply that the measured ΔA515-560nm trace on a long time scale reflects the interplay of the ΔΨ components. Simulations also show that partitioning of proton motive force (pmf) to ΔΨb and ΔpH components is sensitive to the stoichiometric ratio of H(+)/ATP, energy barrier for ATP synthesis, ionic strength, buffer capacity and light intensity. Our model shows that high buffer capacity promotes the establishment of ΔΨb, while the formation of pHi minimum is not 'dissipated' but 'postponed' until it reaches the same level as that for low buffer capacity. Under physiologically optimal conditions, the output of the model shows that at steady state in light, the ΔpH component is the main contributor to pmf to drive ATP synthesis while a low ΔΨb persists energizing the membrane. Our model predicts 11mV as the resting electric potential difference across the thylakoid membrane in dark. We suggest that the model presented in this work can be integrated as a module into a more comprehensive model of oxygenic photosynthesis.
- MeSH
- biologické modely * MeSH
- časové faktory MeSH
- koncentrace vodíkových iontů účinky záření MeSH
- listy rostlin metabolismus účinky záření MeSH
- membránové potenciály účinky záření MeSH
- počítačová simulace MeSH
- protonmotorická síla účinky záření MeSH
- protony MeSH
- pufry MeSH
- světlo * MeSH
- transport elektronů MeSH
- tylakoidy metabolismus účinky záření MeSH
- uhlík metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
The carrier ampholytes-based (CA-based) isoelectric focusing (IEF) experiment evolved from Svensson's closed system IEF (constant spatial current density, absence of convective mixing, counter-balancing electrophoretic and diffusive fluxes yielding a steady state pH gradient) to the contemporary open system IEF (absence of convective mixing, large cross-sectional area electrode vessels, lack of counter-balancing electrophoretic- and diffusive fluxes leading to transient pH gradients). Open system IEF currently is described by a two-stage model: In the first stage, a rapid IEF process forms the pH gradient which, in the second stage, is slowly degraded by isotachophoretic processes that move the most acidic and most basic CAs into the electrode vessels. An analysis of the effective mobilities and the effective mobility to conductivity ratios of the anolyte, catholyte, and the CAs indicates that in open system IEF experiments a single process, transient bidirectional isotachophoresis (tbdITP) operates from the moment current is turned on until it is turned off. In tbdITP, the anolyte and catholyte provide the leading ions and the pI 7 CA or the reactive boundary of the counter-migrating H3 O+ and OH- ions serves as the shared terminator. The outcome of the tbdITP process is determined by the ionic mobilities, pKa values, and loaded amounts of all ionic and ionizable components: It is constrained by both the transmitted amount of charge and the migration space available for the leading ions. tbdITP and the resulting pH gradient can never reach steady state with respect to the spatial coordinate of the separation channel.
In modern isoelectric focusing (IEF) systems, where (i) convective mixing is prevented by gels or small cross-sectional area separation channels, (ii) current densities vary spatially due to the presence of electrode vessels with much larger cross-sectional areas than those of the gels or separation channels, and (iii) electrophoretic and diffusive fluxes do not balance each other, stationary, steady-state pH gradients cannot form (open-system IEF). Open-system IEF is currently described as a two-stage process: A rapid IEF process forms the pH gradient from the carrier ampholytes (CAs) in the first stage, then isotachophoretic processes degrade the pH gradient in the second stage as the extreme pI CAs are moved into the electrode vessels where they become diluted. Based on the ratios of the local effective mobilities and the local conductivities ( μLeff(x)$\mu _{\rm{L}}^{{\rm{eff}}}( x )$ / κ(x)$\kappa ( x )$ values) of the anolyte, catholyte, and the CAs, we pointed out in the preceding paper (Vigh G, Gas B, Electrophoresis 2023, 44, 675-88) that in open-system IEF, a single process, transient, bidirectional isotachophoresis (tbdITP) operates from the moment current is turned on. In this paper, we demonstrate some of the operational features of the tbdITP model using the new ITP/IEF version of Simul 6.
- Phosphorus-31 NMR Spectroscopy 78 -- 2.3.1 Identification of Resonances 79 -- 2.3.2 Intracellular pH 10.7.2 Signal Reception 537 -- 10.7.3 Quadrature Detection 539 -- 10.7.4 Dynamic Range 540 -- 10.7.5 Gradient
2nd ed. xxi, 570 s., [8] s. obr. příl. : il. ; 25 cm
-- Heterochromatin Is Highly Organized and Restricts Gene -- Expression 194 -- The Heterochromatic State Ion-Concentration Gradients 601 -- CONTENTS -- xxvi i -- ; n the Plasma Membrane Regulate Cytosolic pH Between Open Odsed States -- Potential in Animal Cells Depends Mainly on K+ l barrels and the K+ Gradient Without Cytokinesis 1002 -- s = Stable State of Cdk Inactivity 1002 -- 1004 -- 1004 -- 1004 1006 -- v Goes Wrong -- CP SELL DIVISION AND CELL GROWTH -- e Cell Division -- 5 Specialized Nondividing State
Sixth edition xxxiv, 1430 stran v různém stránkování : ilustrace (převážně barevné) ; 29 cm
"As the amount of information in biology expands dramatically, it becomes increasingly important for textbooks to distill the vast amount of scientific knowledge into concise principles and enduring concepts. As with previous editions, Molecular Biology of the Cell, Sixth Edition accomplishes this goal with clear writing and beautiful illustrations. The Sixth Edition has been extensively revised and updated with the latest research in the field of cell biology, and it provides an exceptional framework for teaching and learning. The entire illustration program has been greatly enhanced. Protein structures better illustrate structure-function relationships, icons are simpler and more consistent within and between chapters, and micrographs have been refreshed and updated with newer, clearer, or better images. As a new feature, each chapter now contains intriguing open-ended questions highlighting "What We Don't Know," introducing students to challenging areas of future research. Updated end-of-chapter problems reflect new research discussed in the text. Thought-provoking end-of-chapter questions have been expanded to all chapters, including questions on developmental biology, tissues and stem cells, the immune system, and pathogens"--Provided by publisher.
- Konspekt
- Biochemie. Molekulární biologie. Biofyzika
- NLK Obory
- molekulární biologie, molekulární medicína
- NLK Publikační typ
- učebnice vysokých škol
State 50 -- Dissociation Constants of Binding Reactions Reflect -- the Affinity of Interacting Molecules 50 -- Biological Fluids Have Characteristic pH Values 51 -- Hydrogen Ions Are Released by Acids and -- Taken Up by Bases 52 -- Buffers Maintain the pH of Intracellular and -- Extracellular Fluids 52 -- a Reaction Depends on the Activation Energy Necessary to Energize the Reactants into a Transition State Glucose and Na + -- Parietal Cells Acidify the Stomach Contents While Maintaining a Neutral Cytosolic pH
6th ed. xxxvii, 1150 s. : il., tab. ; 29 cm
- MeSH
- biologie buňky MeSH
- molekulární biologie MeSH
- Publikační typ
- monografie MeSH
- Konspekt
- Biochemie. Molekulární biologie. Biofyzika
- NLK Obory
- biologie
- cytologie, klinická cytologie
