Biomedical subjects

M Pasenkiewicz-Gierula

Publications and source records attributed to M Pasenkiewicz-Gierula.

24 records · Page 2Linked to original sources

PubMed1987

Adduct formation between the cupric site of phenylalanine hydroxylase from Chromobacterium violaceum and 6,7-dimethyltetrahydropterin.

The interaction of pterin-dependent phenylalanine hydroxylase from Chromobacterium violaceum with the cofactor analogue 5-deaza-6-methyltetrahydropterin and the cofactor 6,7-dimethyltetrahydropterin (DMPH4) has been investigated by multifrequency electron spin resonance (ESR) spectroscopy. 5-Deaza-6-methyltetrahydropterin, which lacks the N-5 nitrogen present in the pyrazine ring of DMPH4, binds tightly to the cupric form of the enzyme; however, no changes are observed in the ESR parameters of the copper center. In contrast, the binding of DMPH4 (or 6-methyltetrahydropterin) shifts the ESR parameters (g and A) associated with the cupric enzyme. In addition, superhyperfine transitions were resolved and assigned to hyperfine splitting from nitrogen ligands. ESR spectra of the enzyme recorded in the presence of [5-14N]DMPH4 or [5-15N]DMPH4 were computer simulated and found to be consistent with pterin serving as a direct donor ligand to the copper center through the N-5 position.

Binding Sites↗

PubMed1987

Lateral diffusion of lipids in membranes by pulse saturation recovery electron spin resonance.

Short-pulse saturation recovery electron spin resonance methods have been used to measure lateral diffusion of nitroxide-labeled lipids in multilamellar liposomal dispersions. Nitroxides with 14N and 15N isotopes introduced both separately and together were used. Differential equations have been written and solved for complex saturation recovery signals involving several superimposed exponentials. The time constants contain various combinations of the spin-lattice relaxation time (T1e) for both isotopes, Heisenberg exchange rates, and nuclear spin-lattice relaxation times (T1n). Signals of high quality were fitted by Monte Carlo variation of the amplitudes and time constants. The reliability of the approach was tested extensively by verifying that (i) the predicted number of exponentials agreed with the experimental number, (ii) relaxation parameters that were determined were independent of the observed hyperfine transition, (iii) the time constants were independent of saturating pulse length, (iv) T1e and T1n do not change when Heisenberg exchange is changed by varying the concentration, and (v) Heisenberg exchange is indeed proportional to the concentration. It has been established that bimolecular collision rates over a wide range of conditions can be reliably measured using the methodology described here. The methods depend on the favorable match of bimolecular collision rates at micromolar concentrations to nitroxide spin-lattice relaxation probabilities.

Diffusion↗

PubMed1986

Analysis of the ESR spectrum of synthetic dopa melanin.

The 35 GHz ESR spectrum of frozen aqueous suspensions of synthetic melanin from autoxidation of dopa is asymmetric in the pH range 3-12. This asymmetry increases with increasing pH. A detailed computer analysis of second-derivative spectra suggests that the asymmetry is a result of two factors: approximately axial anisotropy of the g-tensor of the radical species; superposition of ESR spectra arising from a total of four radical species. The relative amounts of these individual spectra vary in a pH-dependent manner. Anisotropy of spectra varies with pH, probably due to pH-induced changes in the delocalization of the unpaired electrons. Thus the species present at high pH are suggested to be relatively localized radical anions, while the species detected at low pH are suggested to be protonated forms of the high pH species in which the unpaired electron is more extensively delocalized.

Dihydroxyphenylalanine↗

PubMed1986

Spin-label studies on phosphatidylcholine-cholesterol membranes: effects of alkyl chain length and unsaturation in the fluid phase.

Dynamic properties of phosphatidylcholine-cholesterol membranes in the fluid phase and water accessibility to the membranes have been studied as a function of phospholipid alkyl chain length, saturation, mole fraction of cholesterol, and temperature by using spin and fluorescence labelling methods. The results are the following: (1) The effect of cholesterol on motional freedom of 5-doxyl stearic acid spin label (5-SASL) and 16-doxyl stearic acid spin label (16-SASL) in saturated phosphatidylcholine membrane is significantly larger than the effects of alkyl chain length and introduction of unsaturation in the alkyl chain. (2) Variation of alkyl chain length of saturated phospholipids does not alter the effects of cholesterol except in the case of dilauroylphosphatidylcholine, which possesses the shortest alkyl chains (12 carbons) used in this work. (3) Unsaturation of the alkyl chains greatly reduces the ordering effect of cholesterol at C-5 and C-16 positions although unsaturation alone gives only minor fluidizing effects. (4) Introduction of 30 mol% cholesterol to dimyristoylphosphatidylcholine membranes decreases the lateral diffusion constants of lipids by a factor of four, while it causes only a slight decrease of lateral diffusion in dioleoylphosphatidylcholine membranes. (5) If compared at the same temperature, 5-SASL mobilities plotted as a function of mole fraction of cholesterol in the fluid phases of dimyristoylphosphatidylcholine-, dipalmitoylphosphatidylcholine- and distearoylphosphatidylcholine-cholesterol membranes are similar in wide ranges of temperature (45-82 degrees C) and cholesterol mole fraction (0-50%). (6) In isothermal experiments with saturated phosphatidylcholine membranes, 5-SASL is maximally immobilized at the phase boundary between Regions I and III reported by other workers (Recktenwald, D.J. and McConnell, H.M. (1981) Biochemistry 20, 4505-4510) and becomes more mobile away from the boundary in Regions I and III. (7) 5-SASL in unsaturated phosphatidylcholine membranes showed a gradual monotonic immobilization with increase of cholesterol mole fraction without showing any maximum in the range of cholesterol fractions studied. (8) By rigorously determining rigid-limit magnetic parameters of cholestane spin labels in membranes from Q-band second-derivative ESR spectra to monitor the dielectric environment around the nitroxide radical, it is concluded that cholesterol incorporation increases water accessibility in the hydrophilic loci of the membrane. In contrast, 12-(9-anthroyloxy)stearic acid fluorescence showed that water accessibility is decreased in the hydrophobic loci of the membrane.

Cholesterol↗

PubMedDate not supplied

Molecular dynamics of a vasopressin V2 receptor in a phospholipid bilayer membrane.

Molecular dynamics simulations were carried out for a V2 receptor (V2R) model embedded in a dimyristoylphosphatidylcholine (DMPC) bilayer. Both free and ligand-bound states of V2R were modeled. Our initial V2R model was obtained using a rule-based automated method for GPCR modeling and refined using constrained simulated annealing in vacuo. The docking site of the native vasopressin ligand was selected and justified upon consideration of ligand-receptor interactions and structure-activity data. The primary purpose of this work was to investigate the usefulness of MD simulation of an integral membrane protein like a GPCR receptor, upon inclusion of a carefully parameterized surrounding lipid membrane and water. Physical properties of the system were evaluated and compared with the fully hydrated pure DMPC bilayer membrane. The solvation interactions, individual lipid-protein interaction and fluctuations of the protein, the lipid, and water were analyzed in detail. As expected, the membrane-spanning helices of the protein fluctuate less than the peripheral loops do. The protein appears to disturb the local lipid structure. Simulations were carried out using AMBER 4.1 package upon constant number-pressure-temperature (NPT) conditions on massively parallel computers Cray T3E and IBM SP2.

Animals↗

PubMedDate not supplied

Comparative investigations of hydroxyamine carane derivative and its R,S-diastereoisomers with strong local anesthetic activity.

Our previously conducted pharmacological screening led as to the discovery of the strong local anesthetic activity of the compound designated as KP-23. Earlier crystallographic studies revealed that the compound KP-23 crystallized in diastereoisomeric form in lowest symmetry. The aim of these comparative investigations was to evaluate anesthetic activity of KP-23 and its R,S-diastereoisomers, which were synthesized at the Institute of Organic Chemistry, Biochemistry and Biotechnology, Wroclaw University of Technology.

Anesthetics, Local↗