Side-chain conformers to allow conversion from normal to isoaspartate in age-related proteins and peptides.

Dβ (or D-iso)- and Lβ- (or iso)- aspartyl (Asp) residues are accumulated in aged lens crystallins and amyloid beta (Aβ) proteins, respectively, as a result of spontaneous, nonenzymatic isomerization of normal Lα-Asp. To explore why such uncommon Asp isomers are accumulated, the stability of Lα-, Lβ-, and Dβ-Asp was compared in view of the staggered side-chain conformers. By using cylindrin (KVKVLGDVIEV) from αB-crystallin and Aβ17-25 (LVFFAED)VG) containing Asp isomers, the vicinal spin-spin coupling constants of Asp H-H and H-H were quantified by high-resolution solution H NMR.

Solution NMR to Quantify Mobility in Membranes: Diffusion, Protrusion, and Drug Transport Processes.

Lipid bilayer membranes are soft, fluid, and dynamic architecture where molecules are constantly moving and thermally fluctuating under physiological conditions. In this review, a strategy to quantify molecular dynamics in membranes is introduced by utilizing solution-state NMR spectroscopy as a versatile, noninvasive technique. The dynamics involves lateral diffusion and protrusion motion, in parallel and vertical direction to the membrane surface.

Evaluation of the diameter of the distal radial artery at the anatomical snuff box using ultrasound in Japanese patients.

Catheter angioplasty or angiography via the distal access point of the radial artery (dRA), located at the anatomical snuff box, is a less invasive strategy for coronary intervention attracting considerable attention. Determining the diameter of the dRA is necessary to minimize the risk of artery occlusion and safely perform catheter intervention. This was a retrospective observational study including patients who underwent coronary angiography or coronary intervention at Aomori Kyoritsu Hospital, Aomori, Japan, between February 2018 and August 2018.

Glycosaminoglycan Binding and Non-Endocytic Membrane Translocation of Cell-Permeable Octaarginine Monitored by Real-Time In-Cell NMR Spectroscopy.

Glycosaminoglycans (GAGs), which are covalently-linked membrane proteins at the cell surface have recently been suggested to involve in not only endocytic cellular uptake but also non-endocytic direct cell membrane translocation of arginine-rich cell-penetrating peptides (CPPs). However, in-situ comprehensive observation and the quantitative analysis of the direct membrane translocation processes are challenging, and the mechanism therefore remains still unresolved. In this work, real-time in-cell NMR spectroscopy was applied to investigate the direct membrane translocation of octaarginine (R8) into living cells.

Kinetics of the competitive reactions of isomerization and peptide bond cleavage at l-α- and d-β-aspartyl residues in an αA-crystallin fragment.

d-β-aspartyl (Asp) residue has been found in a living body such as aged lens crystallin, although l-α-amino acids are constituents in natural proteins. Isomerization from l-α- to d-β-Asp probably modulates structures to affect biochemical reactions. At Asp residue, isomerization and peptide bond cleavage compete with each other.

D-β-aspartyl residue exhibiting uncommon high resistance to spontaneous peptide bond cleavage.

Although L-amino acids were selected as main constituents of peptides and proteins during chemical evolution, D-aspartyl (Asp) residue is found in a variety of living tissues. In particular, D-β-Asp is thought to be stable than any other Asp isomers, and this could be a reason for gradual accumulation in abnormal proteins and peptides to modify their structures and functions. It is predicted that D-β-Asp shows high resistance to biomolecular reactions.

Uptake of sevoflurane limited by the presence of cholesterol in the lipid bilayer membrane: a multinuclear nuclear magnetic resonance study.

The effect of cholesterol on the uptake of a fluorinated general anesthetic, sevoflurane (SF, fluoromethyl 2,2,2-trifluoro-1-[trifluoromethyl]ethyl ether) was studied by multinuclear, high-resolution nuclear magnetic resonance (NMR) spectroscopy in combination with a pulsed-field gradient technique. Using large unilamellar vesicles of egg phosphatidylcholine/egg phosphatidylglycerol/cholesterol as model fluid cell membranes, the (19)F and (1)H NMR chemical shifts, longitudinal relaxation times (T1), and diffusion coefficients (D(eff)) were systematically analyzed to quantify the modulation of SF uptake to the lipid membrane by cholesterol. All NMR parameters (chemical shift, T1, and D(eff)) showed that SF uptake is limited by the presence of cholesterol in the membrane.

Staggered side-chain conformers of aspartyl residues prerequisite to transformation from L-α- to D-β-aspartate 58 in human-lens αA-crystallin fragment.

D-β-aspartyl (Asp) residues are found in aged human-lens αA-crystallin. To explore why the uncommon D-β-Asp is accumulated, the stability of L-α-, D-α-, and D-β-Asp residues is compared in view of the staggered side-chain conformers. By using αA-crystallin fragment, T(55)VLD(58)SGISEVR(65), composed of Asp58 isomers, the vicinal spin-spin coupling constants of Asp58 Hα-Hβ1 and Hα-Hβ2 are quantified by high-resolution solution NMR.

Lateral diffusion of lipids separated from rotational and translational diffusion of a fluid large unilamellar vesicle.

A new method to separate lateral diffusion of lipids in spherical large unilamellar vesicles from the rotational and the translational diffusion of the vesicle as a whole is proposed. The lateral diffusion coefficient DL is obtained as a time-dependent part of the observed diffusion coefficient in vesicles of 800-nm diameters, by systematically changing the diffusion time interval of the high-field-gradient NMR measurement. Although the lipid is in a confined space, the DL of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine is (1.

Kinetics of binding and diffusivity of leucine-enkephalin in large unilamellar vesicle by pulsed-field-gradient H NMR .

The kinetics of binding, the diffusivity, and the binding amount of a neuropeptide, leucine-enkephalin (L-Enk) to lipid bilayer membranes are quantified by pulsed-field-gradient (PFG) H NMR . The peptide signal is analyzed by the solution of the Bloch equation with exchange terms in the presence of large unilamellar vesicles (LUVs) as confined, but fluid model cell membranes. Even in the case that the membrane-bound and the free states of L-Enk cannot be distinguished in the one-dimensional NMR spectrum, the PFG technique unveils the bound component of L-Enk after the preferential decay of the free component at the high field gradient.

Binding of hydrophobic fluorinated bisphenol A to large unilamellar vesicles of egg phosphatidylcholine.

The kinetics of membrane binding and dissociation of fluorinated bisphenol A (FBPA, (CF(3))(2)C(C(6)H(4)OH)(2)) is quantified by 1D (19)F NMR spectra in situ. Although the bound and free components are in fast exchange, the rate constants and bound fraction is nonetheless determined from an analysis of the spectra. The analysis relies on the expression of 1D NMR signal intensity by a set of Bloch equations with exchange terms.

Drug binding and mobility relating to the thermal fluctuation in fluid lipid membranes.

Drug binding and mobility in fluid lipid bilayer membranes are quantified in situ by using the multinuclear solution NMR combined with the pulsed-field-gradient technique. One-dimensional and pulsed-field-gradient (19)F and (1)H NMR signals of an anticancer drug, 5-fluorouracil (5FU) are analyzed at 283-313 K in the presence of large unilamellar vesicles (LUVs) of egg phosphatidylcholine (EPC) as model cell membranes. The simultaneous observation of the membrane-bound and free 5FU signals enables to quantify in what amount of 5FU is bound to the membrane and how fast 5FU is moving within the membrane in relation to the thermal fluctuation of the soft, fluid environment.

Cholesterol location and orientation in aqueous suspension of large unilamellar vesicles of phospholipid revealed by intermolecular nuclear overhauser effect.

The locational and orientational structure and the dynamics of cholesterol in the bilayer membrane were studied by using the solution-state NMR. The intermolecular nuclear Overhauser effect (NOE) was analyzed for large unilamellar vesicles (100 nm in diameter) composed of dimyristoylphosphatidylcholine (DMPC) and cholesterol at cholesterol concentrations of 9-33 mol %. The DMPC headgroups show (1)H-{(1)H}-NOEs with the methyl groups at the hydrophobic terminals of both cholesterol and DMPC, illustrating the significant fluctuation of the bilayer membrane in the vertical (bilayer normal) direction.

Dynamic and 2D NMR studies on hydrogen-bonding aggregates of cholesterol in low-polarity organic solvents.

Self-diffusion coefficients (D) are measured for normal (nondeuterated) and deuterated cholesterol-d(6) (C26 and C27 methyl groups deuterated) in 1-octanol, chloroform, and cyclohexane at concentrations of 1-700 mM by varying the impurity water concentration (>2 mM) and temperature (30-50 degrees C). The pulsed field gradient spin-echo (PGSE) (1)H and (2)H NMR were used, respectively, at 600 and 92 MHz. At 30 degrees C, the hydrodynamic radius (R) obtained at 20 mM from the D value and solvent viscosity is 5.

Limited slowdown of endocrine-disruptor diffusion in confined fluid lipid membranes.

Self-diffusion rates of lipids and trapped bisphenol A (BPA) are determined in various sizes of confined but fluid membranes by high-field-gradient NMR at 600 MHz. Micelles and vesicles of 3- to 400-nm diameters are used as model membranes to get an insight into the molecular diffusion in such soft environments. The slowdown of BPA and lipid motions is leveled off in 100- and 400-nm vesicles, although the hydrodynamic continuum model gives the aggregate motion slowed inversely to the aggregate size.

NMR study on the binding of neuropeptide achatin-I to phospholipid bilayer: the equilibrium, location, and peptide conformation.

Molecular mechanism of the binding of neuropeptide achatin-I (Gly-D-Phe-Ala-Asp) to large unilamellar vesicles of zwitterionic egg-yolk phosphatidylcholine (EPC) was investigated by means of natural-abundance (13)C and high-resolution (of 0.01 Hz order) (1)H NMR spectroscopy. The binding equilibrium was found to be sensitive to the ionization state of the N-terminal NH(3)(+) group in achatin-I; the de-ionization of NH(3)(+) decreases the bound fraction of the peptide from approximately 15% to nearly none.