Replacement of His23 by Cys in a zinc finger of HIV-1 NCp7 led to a change in 1H NMR-derived 3D structure and to a loss of biological activity

AbstractThe nucleocapsid protein NCp7 of human immunodeficiency virus type 1 (HIV-1), which is necessary for the formation of infectious virions, contains two zinc fingers of the Cys-X2-Cys-X4-His-X4-Cys form. To elucidate the importance of this particular motif, well conserved in retroviruses and retroelements, we substituted the histidine residue by a cysteine in the first zinc binding domain 13VKCFNCGKEGHTARNCRA30. The structures of the mutated and native zinc complexed peptides were studied by two-dimensional 600 MHz 1H nuclear magnetic resonance (NMR) in aqueous solution. The nuclear Overhauser effects were used as constraints to determine the solution structures using DIANA software followed by AMBER energy refinement. The results show that native and mutant peptides fold into non-identical three-dimensional structures, probably accounting for the loss of retrovirus infectivity following the His-Cys point mutation

Translocation Of The pAntp Peptide And Its Amphipathic Analogue Ap-2al

The pAntp peptide, corresponding to the third helix of the homeodomain of the Antennapedia protein, enters by a receptor-independent process into eukaryotic cells. The interaction between the pAntp peptide and the phospholipid matrix of the plasma membrane seems to be the first step involved in the translocation mechanism. However, the mechanism by which the peptide translocates through the cell membrane is still not well established. We have investigated the translocation ability of pAntp through a protein-free phospholipid membrane in comparison with a more amphipathic analogue. We show by fluorescence spectroscopy, circular dichroism, NMR spectroscopy, and molecular modeling that pAntp is not sufficiently helically amphipathic to cross a phospholipid membrane of a model system. Due to its primary sequence related to its DNA binding ability in the Antennapedia homeodomain-DNA complex, the pAntp peptide does not belong to the amphipathic alpha-helical peptide family whose members are able to translocate by pore formation

A simple novel approach for detecting blood–brain barrier permeability using GPCR internalization

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Ultrasound localization microscopy to image and assess microvasculature in a rat kidney

Abstract The recent development of ultrasound localization microscopy, where individual microbubbles (contrast agents) are detected and tracked within the vasculature, provides new opportunities for imaging the vasculature of entire organs with a spatial resolution below the diffraction limit. In stationary tissue, recent studies have demonstrated a theoretical resolution on the order of microns. In this work, single microbubbles were localized in vivo in a rat kidney using a dedicated high frame rate imaging sequence. Organ motion was tracked by assuming rigid motion (translation and rotation) and appropriate correction was applied. In contrast to previous work, coherence-based non-linear phase inversion processing was used to reject tissue echoes while maintaining echoes from very slowly moving microbubbles. Blood velocity in the small vessels was estimated by tracking microbubbles, demonstrating the potential of this technique to improve vascular characterization. Previous optical studies of microbubbles in vessels of approximately 20 microns have shown that expansion is constrained, suggesting that microbubble echoes would be difficult to detect in such regions. We therefore utilized the echoes from individual MBs as microscopic sensors of slow flow associated with such vessels and demonstrate that highly correlated, wideband echoes are detected from individual microbubbles in vessels with flow rates below 2 mm/s