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- A Computational Study of the Open and Closed Forms of the N-...A Computational Study of the Open and Closed Forms of the N-Lobe Human Serum Transferrin Apoprotein
Biophysical Journal, Volume 85, Issue 6, 1 December 2003, Pages 3485-3501
David Rinaldo and Martin J. Field
AbstractHuman serum transferrin tightly binds ferric ions in the blood stream but is able to release them in cells by a process involving receptor-mediated endocytosis and decrease in pH. Iron binding and release are accompanied by a large conformation change. In this study, we investigate theoretically the open and closed forms of the N-lobe human serum transferrin apoprotein by performing pK calculations and molecular dynamics and free-energy simulations. In agreement with the hypothesis based on the x-ray crystal structures, our calculations show that there is a shift in the pK values of the lysines forming the when the conformation changes. We argue, however, that simple electrostatic repulsion between the lysines is not sufficient to trigger domain opening and, instead, propose an alternative explanation for the dilysine-trigger effect. Analysis of the molecular dynamics and free-energy results indicate that the open form is more mobile than the closed form and is much more stable at pH 5.3, in large part due to entropic effects. Despite a lower free energy, the dynamics simulation of the open form shows that it is flexible enough to sample conformations that are consistent with iron binding.
Abstract | Full Text | PDF (527 kb) - On the Equivalence Point for Ammonium (De)protonation during...On the Equivalence Point for Ammonium (De)protonation during Its Transport through the AmtB Channel
Biophysical Journal, Volume 92, Issue 12, 15 June 2007, Pages L103-L105
David L. Bostick and Charles L. Brooks
AbstractStructural characterization of the bacterial channel, AmtB, provides a glimpse of how members of its family might control the protonated state of permeant ammonium to allow for its selective passage across the membrane. In a recent study, we employed a combination of simulation techniques that suggested ammonium is deprotonated and reprotonated near dehydrative phenylalanine landmarks (F107 and F31, respectively) during its passage from the periplasm to the cytoplasm. At these landmarks, ammonium is forced to maintain a critical number (∼3) of hydrogen bonds, suggesting that the channel controls ammonium (de)protonation by controlling its coordination/hydration. In the work presented here, a free energy-based analysis of ammonium hydration in dilute aqueous solution indicates, explicitly, that at biological pH, the transition from ammonium () to ammonia (NH) occurs when these species are constrained to donate three hydrogen bonds or less. This result demonstrates the viability of the proposal that AmtB indirectly controls ammonium (de)protonation by directly controlling its hydration.
Abstract | Full Text | PDF (217 kb) - Fourier Transform Infrared Evidence for Early Deprotonation ...Fourier Transform Infrared Evidence for Early Deprotonation of Asp at Alkaline pH in the Photocycle of Bacteriorhodopsin Mutants Containing E194Q
Biophysical Journal, Volume 78, Issue 4, 1 April 2000, Pages 2022-2030
Tzvetana Lazarova, Carolina Sanz, Enric Querol and Esteve Padrós
AbstractThe role of the extracellular Glu side chains of bacteriorhodopsin in the proton transport mechanism has been studied using the single mutants E9Q, E74Q, E194Q, and E204Q; the triple mutant E9Q/E194Q/E204Q; and the quadruple mutant E9Q/E74Q/E194Q/E204Q. Steady-state difference and deconvoluted Fourier transform infrared spectroscopy has been applied to analyze the M- and N-like intermediates in membrane films maintained at a controlled humidity, at 243 and 277K at alkaline pH. The mutants E9Q and E74Q gave spectra similar to that of wild type, whereas E194Q, E9Q/E194Q/E204Q, and E9Q/E74Q/E194Q/E204Q showed at 277K a N-like intermediate with a single negative peak at 1742cm, indicating that Asp and Asp are deprotonated. Under the same conditions E204Q showed a positive peak at 1762cm and a negative peak at 1742cm, revealing the presence of protonated Asp (in an M intermediate environment) and deprotonated Asp. These results indicate that in E194Q-containing mutants, the second increase in the Asp pK is inhibited because of lack of deprotonation of the proton release group. Our data suggest that Glu is the group that controls the pK of Asp.
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Copyright © 1998 The Biophysical Society. All rights reserved.
Biophysical Journal, Volume 74, Issue 6, 2747-2759, 1 June 1998
doi:10.1016/S0006-3495(98)77983-4
The pH-Induced Release of Iron from Transferrin Investigated with a Continuum Electrostatic Model
David A. Lee and Julia M. Goodfellow
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Address reprint requests to Dr. Julia Goodfellow, Department of Crystallography, Birkbeck College, Malet Street, London WC1E 7HX, UK. Tel.: 44-171-631-6833; Fax: 44-171-631-6833.Abstract
A reduction in pH induces the release of iron from transferrin in a process that involves a conformational change in the protein from a closed to an open form. Experimental evidence suggests that there must be changes in the protonation states of certain, as yet not clearly identified, residues in the protein accompanying this conformational change. Such changes in protonation states of residues and the consequent changes in electrostatic interactions are assumed to play a large part in the mechanism of release of iron from transferrin. Using the x-ray crystal structures of human ferri- and apo-lactoferrin, we calculated the pKa values of the titratable residues in both the closed (iron-loaded) and open (iron-free) conformations with a continuum electrostatic model. With the knowledge of a residue’s pKa value, its most probable protonation state at any specified pH may be determined. The preliminary results presented here are in good agreement with the experimental observation that the binding of ferric iron and the synergistic anion bicarbonate/carbonate results in the release of approximately three H+ ions. It is suggested that the release of these three H+ ions may be accounted for, in most part, by the deprotonation of the bicarbonate and residues Tyr-92, Lys-243, Lys-282, and Lys-285 together with the protonation of residues Asp-217 and Lys-277.
