Biomedical subjects

Anne S Ulrich

Publications and source records attributed to Anne S Ulrich.

23 records · Page 2Linked to original sources

PubMed2003

Susceptibility corrections in solid-state NMR experiments with oriented membrane samples. Part I: applications.

Chemical shift referencing of solid-state NMR experiments on oriented membranes has to compensate for bulk magnetic susceptibility effects that are associated with the non-spherical sample shape, as described in the accompanying paper [J. Magn. Reson. 164 (2003) 115-127]. The resulting frequency deviations can be on the order of 10 ppm, which is serious for nuclei with a narrow chemical shift anisotropy such as 1H or 13C, and in some cases even 19F. Two referencing schemes are proposed here to compensate for these effects: A flat (0.4 mm) glass container with an isotropic reference molecule dissolved in a thin film of liquid is stacked on top of the oriented membrane sample. Alternatively, the intrinsic proton signal of the hydrated lipid can be used for chemical shift referencing. Further aspects related to magnetic susceptibility are discussed, such as air gaps in susceptibility-matched probeheads, the benefits of shimming, and limitations in the accuracy of orientational constraints. A biological application is illustrated by a series of experiments on the antimicrobial peptide PGLa, aimed at understanding its concentration-dependent membranolytic effect. To address a wide range of molar peptide/lipid ratios between 1:3000 and 1:8, multilayers of hydrated DMPC containing a 19F-labeled peptide were oriented between stacked glass plates. Maintaining an approximately constant amount of peptide gives rise to thick samples (18 plates) at low, and thin samples (3 plates) at high peptide/lipid ratio. Accurate referencing was critical to reveal a small but significant change over 5 ppm in the anisotropic chemical shift of the 19F label on the peptide, indicative of a change in the orientation and/or dynamics of PGLa in the membrane.

Anisotropy↗

PubMed2003

Susceptibility corrections in solid state NMR experiments with oriented membrane samples. Part II: theory.

In solid state NMR analysis of oriented biomembranes the samples typically have the shape of a rectangular block, formed by stacking a number of glass slides coated with the membranes under investigation. Reference material may be provided internally in the volume of the block or as an external layer on its surface, as described in the accompanying paper [J. Magn. Reson. 164 (2003) 104-114]. The demagnetizing field resulting in such non-spheroidal samples is inhomogeneous. It shifts and broadens the NMR lines of both the sample and of the reference, as compared to the ideal of a spherical sample. The magnitude of these effects is typically of the order of a few ppm. To determine the necessary corrections, a general analysis is presented here for the demagnetizing field of a layered sample of rectangular block geometry, with the normal of the layers parallel to the main field or tilted about an axis of the block. The correction to the line position of the block sample is found to be approximately equal to that of the spheroid which can be inscribed into the block, and for which the correction is well known. For an external reference layer, placed on top of the block, the correction can be found by the same approximation, invoking a simple mirror concept. The layered structure of the block can be accounted for by using an average magnetic susceptibility. Sample and support materials contribute to that average according to their volume filling factors. If the sample material is anisotropic at the molecular level, as e.g. lipid bilayers are, the resulting anisotropy of the block is reduced by the filling factor of the sample material.

Anisotropy↗

PubMed2002

Heterogeneous structure of silk fibers from Bombyx mori resolved by 13C solid-state NMR spectroscopy.

The molecular conformation of silk fibrion is characterized by solid-state 13C NMR before spinning (silk I structure) and after spinning (silk II structure). We compare native silk fibers with the quasi-crystalline Cp-fraction and a synthetic model peptide (Ala-Gly)15, both of which can be converted either into silk I by dialysis from 9 M LiBr or into silk II by treatment with formic acid. Our results demonstrate that silk II fibers are intrinsically heterogeneous, consisting of beta-sheets, distorted beta-turns, and distorted beta-sheets. This higher-order heterogeneity is revealed by the 13C-NMR Cbeta-peak of Ala, indicating that the Ala side chains are stacked partially in parallel and partially face-to-face, at a ratio of 1:2.

Animals↗

PubMed2002

Solid-state 19F-NMR analysis of 19F-labeled tryptophan in gramicidin A in oriented membranes.

The response of membrane-associated peptides toward the lipid environment or other binding partners can be monitored by solid-state NMR of suitably labeled side chains. Tryptophan is a prominent amino acid in transmembrane helices, and its (19)F-labeled analogues are generally biocompatible and cause little structural perturbation. Hence, we use 5F-Trp as a highly sensitive NMR probe to monitor the conformation and dynamics of the indole ring. To establish this (19)F-NMR strategy, gramicidin A was labeled with 5F-Trp in position 13 or 15, whose chi(1)/chi(2) torsion angles are known from previous (2)H-NMR studies. First, the alignment of the (19)F chemical shift anisotropy tensor within the membrane was deduced by lineshape analysis of oriented samples. Next, the three principal axes of the (19)F chemical shift anisotropy tensor were assigned within the molecular frame of the indole ring. Finally, determination of chi(1)/chi(2) for 5F-Trp in the lipid gel phase showed that the side chain alignment differs by up to 20 degrees from its known conformation in the liquid crystalline state. The sensitivity gain of (19)F-NMR and the reduction in the amount of material was at least 10-fold compared with previous (2)H-NMR studies on the same system and 100-fold compared with (15)N-NMR.

Cell Membrane↗

PubMed2002

Biophysical aspects of using liposomes as delivery vehicles.

Liposomes are used as biocompatible carriers of drugs, peptides, proteins, plasmic DNA, antisense oligonucleotides or ribozymes, for pharmaceutical, cosmetic, and biochemical purposes. The enormous versatility in particle size and in the physical parameters of the lipids affords an attractive potential for constructing tailor-made vehicles for a wide range of applications. Some of the recent literature will be reviewed here and presented from a biophysical point of view, thus providing a background for the more specialized articles in this special issue on liposome technology. Different properties (size, colloidal behavior, phase transitions, and polymorphism) of diverse lipid formulations (liposomes, lipoplexes, cubic phases, emulsions, and solid lipid nanoparticles) for distinct applications (parenteral, transdermal, pulmonary, and oral administration) will be rationalized in terms of common structural, thermodynamic and kinetic parameters of the lipids. This general biophysical basis helps to understand pharmaceutically relevant aspects such as liposome stability during storage and towards serum, the biodistribution and specific targeting of cargo, and how to trigger drug release and membrane fusion. Methods for the preparation and characterization of liposomal formulations in vitro will be outlined, too.

Animals↗