20 research outputs found
Hemoglobin is an oxygen-dependent glutathione buffer adapting the intracellular reduced glutathione levels to oxygen availability
Fast changes in environmental oxygen availability translate into shifts in mitochondrial free radical production. An increase in intraerythrocytic reduced glutathione (GSH) during deoxygenation would support the detoxification of exogenous oxidants released into the circulation from hypoxic peripheral tissues. Although reported, the mechanism behind this acute oxygen-dependent regulation of GSH in red blood cells remains unknown. This study explores the role of hemoglobin (Hb) in the oxygen-dependent modulation of GSH levels in red blood cells. We have demonstrated that a decrease in Hb O2 saturation to 50% or less observed in healthy humans while at high altitude, or in red blood cell suspensions results in rising of the intraerythrocytic GSH level that is proportional to the reduction in Hb O2 saturation. This effect was not caused by the stimulation of GSH de novo synthesis or its release during deglutathionylation of Hb's cysteines. Using isothermal titration calorimetry and in silico modeling, we observed the non-covalent binding of four molecules of GSH to oxy-Hb and the release of two of them upon deoxygenation. Localization of the GSH binding sites within the Hb molecule was identified. Oxygen-dependent binding of GSH to oxy-Hb and its release upon deoxygenation occurred reciprocally to the binding and release of 2,3-bisphosphoglycerate. Furthermore, noncovalent binding of GSH to Hb moderately increased Hb oxygen affinity. Taken together, our findings have identified an adaptive mechanism by which red blood cells may provide an advanced antioxidant defense to respond to oxidative challenges immediately upon deoxygenation
Lactose binding to human galectin-7 (p53-induced gene 1) induces long-range effects through the protein resulting in increased dimer stability and evidence for positive cooperativity.
16 pags, 11 figs, 3 tabs. -- Supplementary data for this article are available online at: http://glycob.oxfordjournals.org/lookup/suppl/doi:10.1093/glycob/cwt005/-/DC1The product of p53-induced gene 1 is a member of the galectin family, i.e., galectin-7 (Gal-7). To move beyond structural data by X-ray diffraction, we initiated the study of the lectin by nuclear magnetic resonance (NMR) and circular dichroism spectroscopies, and molecular dynamics (MD) simulations. In concert, our results indicate that lactose binding to human Gal-7 induces long-range effects (minor conformational shifts and changes in structural dynamics) throughout the protein that result in stabilization of the dimer state, with evidence for positive cooperativity. Monte Carlo fits of 15N-Gal-7 HSQC titrations with lactose using a two-site model yield K1 = 0.9 ± 0.6 Ă 103 Mâ1 and K2 = 3.4 ± 0.8 Ă 103 Mâ1. Ligand binding-induced stabilization of the Gal-7 dimer was supported by several lines of evidence: MD-based calculations of interaction energies between ligand-loaded and ligand-free states, gel filtration data and hetero-FRET spectroscopy that indicate a highly reduced tendency for dimer dissociation in the presence of lactose, CD-based thermal denaturation showing that the transition temperature of the lectin is significantly increased in the presence of lactose, and saturation transfer difference (STD) NMR using a molecular probe of the monomer state whose presence is diminished in the presence of lactose. MD simulations with the half-loaded ligand-bound state also provided insight into how allosteric signaling may occur. Overall, our results reveal long-range effects on Gal-7 structure and dynamics, which factor into entropic contributions to ligand binding and allow further comparisons with other members of the galectin family.This work was supported by a research grant from the National Institutes of Health (CA 096090 to K.H.M.), the RAS program âMolecular and Cellular Biologyâ RFBR grant (No. 12 04 31360 to E.E.), the European Union Seventh Framework Programme (FP7/2007-2013) under grant agreement no 260600 (âGlycoHITâ), grants CTQ2009-08536 and BFU2009-10052 and a FPI PhD fellowship to M.A.B. from the Spanish Ministry of Science and Innovation and the CIBER of Respiratory Diseases (CIBERES), an initiative from the Spanish Institute of Health Carlos III (ISCIII). E.E. was supported by a Travel Grant from the Minnesota Supercomputing Institute (University of Minnesota) during her stay in the research lab of Prof. K.H. Mayo. I.N. was supported in the Mayo lab by National Institutes of Health Hematology Training Grant (HL 07062
Styrylmalonates as an Alternative to DonorâAcceptor Cyclopropanes in the Reactions with Aldehydes: A Route to 5,6-Dihydropyran-2-ones
A new
strategy for modifying the reactivity of donorâacceptor
cyclopropanes (DAC) has been suggested. It involves the use of isomeric
styrylmalonates as alternative sources of reactive intermediates.
The efficiency of the approach has been demonstrated in reactions
with aromatic aldehydes. As a result, a new process for construction
of the 5,6-dihydroÂpyran-2-one skeleton has been developed. It
efficiently occurs with high diastereoselectivity in the presence
of BF<sub>3</sub>·Et<sub>2</sub>O; the products can be easily
isolated by crystallization. The subsequent use of the resulting dihydropyranones
in syntheses providing convenient access to various classes of compounds
with broad molecular diversity has been demonstrated
Styrylmalonates as an Alternative to DonorâAcceptor Cyclopropanes in the Reactions with Aldehydes: A Route to 5,6-Dihydropyran-2-ones
A new
strategy for modifying the reactivity of donorâacceptor
cyclopropanes (DAC) has been suggested. It involves the use of isomeric
styrylmalonates as alternative sources of reactive intermediates.
The efficiency of the approach has been demonstrated in reactions
with aromatic aldehydes. As a result, a new process for construction
of the 5,6-dihydroÂpyran-2-one skeleton has been developed. It
efficiently occurs with high diastereoselectivity in the presence
of BF<sub>3</sub>·Et<sub>2</sub>O; the products can be easily
isolated by crystallization. The subsequent use of the resulting dihydropyranones
in syntheses providing convenient access to various classes of compounds
with broad molecular diversity has been demonstrated