Thiol-ene reaction: a versatile tool in site-specific labelling of proteins with chemically inert tags for paramagnetic NMR

Site-specific tagging of proteins with paramagnetic lanthanides generates valuable long-range structure restraints for structural biology by NMR spectroscopy. We show that the thiol-ene addition reaction offers a powerful tool for tagging proteins in a chemically stable manner with very small lanthanide tags.     

Thermodiffusion motion of electrically charged nanoparticles

The present work deals with experimental studies to examine the theoretical model of thermodiffusion of electrically charged nanoparticles. Three different ionic magnetic colloid samples have been synthesized and profoundly analyzed. The theoretical model is a classical one, based on the calculation of the temperature and the electric potential distribution around nanoparticles. The discrepancy between experimental data and theory turns out not to exceed 20%. We focus on applying different approximations between calculated electrical double layer in the theoretical model and experimental determination of the surface charge density of colloidal particles. We assume this is the main reason for obtained discrepancy.     

4,4'-dithiobisdipicolinic acid: a small and convenient lanthanide binding tag for protein NMR spectroscopy

Pseudocontact shifts (PCS) from paramagnetic lanthanide ions present powerful long-range structure restraints for studies of proteins by nuclear magnetic resonance spectroscopy. To elicit PCSs, the lanthanide must be attached site-specifically to the target protein. In addition, it needs to be attached rigidly to avoid averaging of the PCSs due to mobility with respect to the protein and it must not interfere with the function of the protein. Here, we present a dipicolinic acid reagent that spontaneously forms a disulfide bond with thiol groups of accessible cysteine residues. A minimal number of rotatable bonds between the cysteine side chain and the tag helps to minimise mobility. Combined with the small size of the tag and quantitative tagging yields, these features make it a highly attractive tool for generating structure restraints by paramagnetic lanthanides.     

Fixed space of positive trace-preserving super-operators

We examine the fixed space of positive trace-preserving super-operators. We describe a specific structure that this space must have and what the projection onto it must look like. We show how these results, in turn, lead to an alternative proof of the complete characterization of the fixed space of completely positive trace-preserving super-operators.     

A kilohertz frame rate cinemagraphic PIV system for laboratory-scale turbulent and unsteady flows

A kilohertz frame rate cinemagraphic particle image velocimetry (PIV) system has been developed for acquiring time-resolved image sequences of laboratory-scale gas and liquid-phase turbulent flows. Up to 8000 instantaneous PIV images per second are obtained, with sequence lengths exceeding 4000 images. The two-frame cross-correlation method employed precludes directional ambiguity and has a higher signal-to-noise ratio than single-frame autocorrelation or cross-correlation methods, facilitating acquisition of long uninterrupted sequences of valid PIV images. Low and high velocities can be measured simultaneously with similar accuracy by adaptively cross-correlating images with the appropriate time delay. Seed particle illumination is provided by two frequency-doubled Nd:YAG lasers producing Q-switched pulses at the camera frame rate. PIV images are acquired using a 16 mm high-speed rotating prism camera. Frame-to-frame registration is accomplished by imaging two pairs of crossed lines onto each frame and aligning the digitized image sequence to these markers using image processing algorithms. No flow disturbance is created by the markers because only their image is projected to the PIV imaging plane, with the physical projection device residing outside the flow field. The frame-to-frame alignment uncertainty contributes 2% to the overall velocity measurement uncertainty, which is otherwise comparable to similar film-based PIV methods. Received: 11 July 2000 / Accepted: 21 June 2001 Published online: 29 November 2001     

Engineering of a bis-chelator motif into a protein α-helix for rigid lanthanide binding and paramagnetic NMR spectroscopy

Attachment of two nitrilotriacetic acid-based ligands to a protein α-helix in an i, i + 4 configuration produces an octadentate chelating motif that is able to bind paramagnetic lanthanide ions rigidly and with high affinity, leading to large pseudocontact shifts and residual dipolar couplings in the NMR spectrum.