Nanoscale building blocks are individually exceptionally strong because they are close to ideal, defect-free materials. It is, however, difficult to retain the ideal properties in macroscale composites. Bottom-up assembly of a clay/polymer nanocomposite allowed for the preparation of a homogeneous, optically transparent material with planar orientation of the alumosilicate nanosheets. The stiffness and tensile strength of these multilayer composites are one order of magnitude greater than those of analogous nanocomposites at a processing temperature that is much lower than those of ceramic or polymer materials with similar characteristics. A high level of ordering of the nanoscale building blocks, combined with dense covalent and hydrogen bonding and stiffening of the polymer chains, leads to highly effective load transfer between nanosheets and the polymer.

Chemical exchange saturation transfer (CEST) is a magnetization transfer (MT) technique to indirectly detect pools of exchangeable protons through the water signal. CEST MRI has focused predominantly on signals from exchangeable protons downfield (higher frequency) from water in the CEST spectrum. Low power radiofrequency (RF) pulses can slowly saturate protons with minimal interference of conventional semi-solid based MT contrast (MTC). When doing so, saturation-transfer signals are revealed upfield from water, which is the frequency range of non-exchangeable aliphatic and olefinic protons. The visibility of such signals indicates the presence of a relayed transfer mechanism to the water signal, while their finite width reflects that these signals are likely due to mobile solutes. It is shown here in protein phantoms and the human brain that these signals build up slower than conventional CEST, at a rate typical for intramolecular nuclear Overhauser enhancement (NOE) effects in mobile macromolecules such as proteins/peptides and lipids. These NOE-based saturation transfer signals show a pH dependence, suggesting that this process is the inverse of the well-known exchange-relayed NOEs in high resolution NMR protein studies, thus an relayed-NOE CEST process. When studying 6 normal volunteers with a low-power pulsed CEST approach, the relayed-NOE CEST effect was about twice as large as the CEST effects downfield and larger in white matter than gray matter. This NOE contrast upfield from water provides a way to study mobile macromolecules in tissue. First data on a tumor patient show reduction in both relayed NOE and CEST amide proton signals leading to an increase in magnetization transfer ratio asymmetry, providing insight into previously reported amide proton transfer (APT) effects in tumors.

Magnetization Transfer Contrast (MTC) and Chemical Exchange Saturation Transfer (CEST) experiments measure the transfer of magnetization from molecular protons to the solvent water protons, an effect that becomes apparent as an MRI signal loss ("saturation"). This allows molecular information to be accessed with the enhanced sensitivity of MRI. In analogy to Magnetic Resonance Spectroscopy (MRS), these saturation data are presented as a function of the chemical shift of participating proton groups, e.g. OH, NH, NH, which is called a Z-spectrum. In tissue, these Z-spectra contain the convolution of multiple saturation transfer effects, including nuclear Overhauser enhancements (NOEs) and chemical exchange contributions from protons in semi-solid and mobile macromolecules or tissue metabolites. As a consequence, their appearance depends on the magnetic field strength (B) and pulse sequence parameters such as B strength, pulse shape and length, and interpulse delay, which presents a major problem for quantification and reproducibility of MTC and CEST effects. The use of higher B can bring several advantages. In addition to higher detection sensitivity (signal-to-noise ratio, SNR), both MTC and CEST studies benefit from longer water T allowing the saturation transferred to water to be retained longer. While MTC studies are non-specific at any field strength, CEST specificity is expected to increase at higher field because of a larger chemical shift dispersion of the resonances of interest (similar to MRS). In addition, shifting to a slower exchange regime at higher B facilitates improved detection of the guanidinium protons of creatine and the inherently broad resonances of the amine protons in glutamate and the hydroxyl protons in myoinositol, glycogen, and glucosaminoglycans. Finally, due to the higher mobility of the contributing protons in CEST versus MTC, many new pulse sequences can be designed to more specifically edit for CEST signals and to remove MTC contributions.

Disruption of the cellular membrane by the amyloidogenic peptide IAPP (or amylin) has been implicated in β-cell death during type 2 diabetes. While the structure of the mostly inert fibrillar form of IAPP has been investigated, the structural details of the highly toxic prefibrillar membrane-bound states of IAPP have been elusive. A recent study showed that a fragment of IAPP (residues 1-19) induces membrane disruption to a similar extent as the full-length peptide. However, unlike the fulllength IAPP peptide, IAPP 1-19 is conformationally stable in an α-helical conformation when bound to the membrane. In vivo and in vitro measurements of membrane disruption indicate the rat version of IAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] , despite differing from hIAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] by the single substitution of Arg18 for His18, is significantly less toxic than hIAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] , in agreement with the low toxicity of the full-length rat IAPP peptide. To investigate the origin of this difference at the atomic level, we have solved the structures of the human and rat IAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] peptides in DPC micelles. While both rat and human IAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] fold into similar mostly α-helical structures in micelles, paramagnetic quenching NMR experiments indicate a significant difference in the membrane orientation of hIAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] and rIAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] . At pH 7.3, the more toxic hIAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] peptide is buried deeper within the micelle, while the less toxic rIAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] peptide is located at the surface of the micelle. Deprotonating H18 in hIAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] reorients the peptide to the surface of the micelle. This change in orientation is in agreement with the significantly reduced ability of hIAPP [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] to cause membrane disruption at pH 6.0. This difference in peptide topology in the membrane may correspond to similar topology differences for the fulllength human and rat IAPP peptides, with the toxic human IAPP peptide adopting a transmembrane orientation and the nontoxic rat IAPP peptide bound to the surface of the membrane.Islet amyloid polypeptide (IAPP, 1 also known as amylin) is one of an increasing number of proteins in which the propensity to form a misfolded state is correlated with pathologies in tissue functioning. Physiologically, IAPP is one of a family of peptides related to calcitonin that act in the control of metabolic functions. Specifically, IAPP acts in concert with insulin to control plasma glucose levels...

Background: cytb 5 modulates catalysis performed by cytsP450, in vivo and in vitro. Results: The structure of full-length cytb 5 was solved by NMR, and the cytP450-binding site on cytb 5 was identified by mutagenesis and NMR. Conclusion: A model of the cytb 5 -cytP450 complex is presented. Addition of a substrate strengthens the cytb 5 -cytP450 interaction. Significance: The cytb 5 -cytP450 complex structure will help unravel the mechanism by which cytb 5 regulates catalysis by cytP450.

Purpose Chemical exchange saturation transfer (CEST) imaging is a new MRI technology allowing the detection of low concentration endogenous cellular proteins and metabolites indirectly through their exchangeable protons. A new technique, variable delay multi-pulse CEST (VDMP-CEST), is proposed to eliminate the need for recording full Z-spectra and performing asymmetry analysis to obtain CEST contrast. Methods The VDMP-CEST scheme involves acquiring images with two (or more) delays between radiofrequency saturation pulses in pulsed CEST, producing a series of CEST images sensitive to the speed of saturation transfer. Subtracting two images or fitting a time series produces CEST and relayed-nuclear Overhauser enhancement CEST maps without effects of direct water saturation and, when using low radiofrequency power, minimal magnetization transfer contrast interference. Results When applied to several model systems (bovine serum albumin, crosslinked bovine serum albumin, l-glutamic acid) and in vivo on healthy rat brain, VDMP-CEST showed sensitivity to slow to intermediate range magnetization transfer processes (rate < 100–150 Hz), such as amide proton transfer and relayed nuclear Overhauser enhancement-CEST. Images for these contrasts could be acquired in short scan times by using a single radiofrequency frequency. Conclusions VDMP-CEST provides an approach to detect CEST effect by sensitizing saturation experiments to slower exchange processes without interference of direct water saturation and without need to acquire Z-spectra and perform asymmetry analysis.

The current study aims to assign and estimate the total creatine (tCr) signal contribution to the Z-spectrum in mouse brain at 11.7 Tesla. Creatine (Cr), phosphocreatine (PCr) and protein phantoms were used to confirm presence of a guanidinium resonance at this field strength. Wild type (WT) and knockout mice with Guanidinoacetate N-Methyltransferase deficiency (GAMT−/−) that have low Cr and PCr concentrations in the brain were used to assign the tCr contribution to the Z-spectrum. To estimate the total guanidinium concentrations, two pools for the Z-spectrum around 2 ppm were assumed: (i) a Lorentzian function representing the guanidinium CEST at 1.95 ppm in the 11.7 T Z-spectrum; (ii) a background signal that can be fitted by a polynomial function. Comparison between the WT and GAMT−/− mice provided strong evidence for three types of contributions to the peak in the Z-spectrum at 1.95 ppm, namely proteins, Cr and PCr, the latter fitted as tCr. A ratio of 20±7% (Protein) and 80±7% tCr was found in brain with 2 μT and 2 s saturation. Based on phantom experiments, the tCr peak was estimated to consist of about 83±5% Cr and 17±5% PCr. Maps for tCr of mouse brain were generated based on the peak at 1.95 ppm after concentration calibration with in vivo MRS.

Islet amyloid polypeptide (IAPP or amylin) is a 37-residue peptide hormone associated with glucose metabolism that is cosecreted with insulin by β-cells in the pancreas. Since human IAPP is a highly amyloidogenic peptide, it has been suggested that the formation of IAPP amyloid fibers is responsible for the death of β-cells during the early stages of type II diabetes. It has been hypothesized that transient membrane-bound α-helical structures of human IAPP are precursors to the formation of these amyloid deposits. On the other hand, rat IAPP forms transient α-helical structures but does not progress further to form amyloid fibrils. To understand the nature of this intermediate state and the difference in toxicity between the rat and human versions of IAPP, we have solved the high-resolution structure of rat IAPP in the membrane-mimicking detergent micelles composed of dodecylphosphocholine. The structure is characterized by a helical region spanning the residues A5 to S23 and a disordered C-terminus. A distortion in the helix is seen at R18 and S19 that may be involved in receptor binding. Paramagnetic quenching NMR experiments indicate that rat IAPP is bound on the surface of the micelle, in agreement with other nontoxic forms of IAPP. A comparison to the detergent-bound structures of other IAPP variants indicates that the N-terminal region may play a crucial role in the self-association and toxicity of IAPP by controlling access to the putative dimerization interface on the hydrophobic face of the amphipathic helix.