Antibacterial Meroterpenoids, Merochlorins G–J from the Marine Bacterium Streptomyces sp.

Four new chlorinated meroterpenoids, merochlorins G−J (1−4), and 10, a dihydronaphthalenedione precursor, along with known merochlorins A (5) and C−F (6−9), were obtained from cultivation of the bacterium strain Streptomyces sp. CNH-189, which was isolated from marine sediment. The planar structures of compounds 1−4 and 10 were elucidated by interpretation of MS, UV, and NMR spectroscopic data. The relative configurations of compounds 1−4 were determined via analysis of nuclear Overhauser effect (NOE) spectroscopic data, after which their absolute configurations were established by comparing the experimental electronic circular dichroism (ECD) spectra of compounds 1−4 to those of previously reported possible enantiomer models and DP4 calculations. Compound 3 displayed strong antibacterial activities against Bacillus subtilis, Kocuria rhizophila, and Staphylococcus aureus, with MIC values of 1, 2, and 2 μg/mL, respectively, whereas compound 1 exhibited weak antibacterial effects on these three strains, with a 16−32 μg/mL MIC value range.

High-affinity tamoxifen analogues retain extensive positional disorder when bound to calmodulin

Using a combination of NMR and fluorescence measurements, we have investigated the structure and dynamics of the complexes formed between calcium-loaded calmodulin (CaM) and the potent breast cancer inhibitor idoxifene, a derivative of tamoxifen. High-affinity binding (Kd∼300 nM) saturates with a 2:1 idoxifene:CaM complex. The complex is an ensemble where each idoxifene molecule is predominantly in the vicinity of one of the two hydrophobic patches of CaM but, in contrast with the lower-affinity antagonists TFP, J-8, and W-7, does not substantially occupy the hydrophobic pocket. At least four idoxifene orientations per domain of CaM are necessary to satisfy the intermolecular nuclear Overhauser effect (NOE) restraints, and this requires that the idoxifene molecules switch rapidly between positions. The CaM molecule is predominantly in the form where the N and C-terminal domains are in close proximity, allowing for the idoxifene molecules to contact both domains simultaneously. Hence, the 2:1 idoxifene:CaM complex illustrates how high-affinity binding occurs without the loss of extensive positional dynamics.

Conformational and Structural characterization of carbohydrates and their interactions studied by NMR

: Carbohydrates, either free or as glycans conjugated with other biomolecules, participate in many essential biological processes. Their apparent simplicity in terms of chemical functionality hides an extraordinary diversity and structural complexity. Deeply deciphering at the atomic level their structures is essential to understand their biological function and activities, but it is still a challenging task in need of complementary approaches and no generalized procedures are available to address the study of such complex, natural glycans. The versatility of Nuclear Magnetic Resonance spectroscopy (NMR) often makes it the preferred choice to study glycans and carbohydrates in solution media. The most basic NMR parameters, namely chemical shifts, coupling constants and nuclear Overhauser effects, allow defining short or repetitive chain sequences and characterize their structures and local geometries either in the free state or when interacting with other biomolecules, rendering additional information on the molecular recognition processes. The increased accessibility to carbohydrate molecules extensively or selectively labeled with 13C boosts the resolution and detail that analyzed glycan structures can reach. In turn, structural information derived from NMR, complemented with molecular modeling and theoretical calculations can also provide dynamic information on the conformational flexibility of carbohydrate structures. Furthermore, using partially oriented media or paramagnetic perturbations, it has been possible to introduce additional long-range observables rendering structural information on longer and branched glycan chains. In this review, we provide examples of these studies and an overview of the recent and most relevant NMR applications in the glycobiology field.

Concentration and Composition Dependent Aggregation of Pluronic- and Poly-(2-Oxazolin)-Efavirenz Formulations in Biorelevant Media

Interactions of intestinal fluids with polymer excipients, drugs and their formulations are not fully understood. Here, diffusion ordered spectroscopy (DOSY) and nuclear Overhauser effect spectroscopy (NOESY), complemented by cryo-TEM were employed to address this. Efavirenz as model drug, the triblock copolymers Pluronic F-127 (PF127) and poly(2-oxazolin) based pMeOx-b-pPrOzi-b-pMeOx (pOx/pOzi) and their respective formulations were studied in simulated fed-state intestinal fluid (FeSSIF). For the individual polymers, the bile interfering nature of PF127 was confirmed and pure pOx/pOzi was newly classified as non-interfering. A different and more complex behaviour was observed if EFV was involved. The formulations showed multi-facetted concentration and composition dependent aggregation. This demonstrates that separate evaluation of polymers or drugs in biorelevant media is not sufficient and their mixtures need to be carefully studied.<br>

Concentration and Composition Dependent Aggregation of Pluronic- and Poly-(2-Oxazolin)-Efavirenz Formulations in Biorelevant Media

Interactions of intestinal fluids with polymer excipients, drugs and their formulations are not fully understood. Here, diffusion ordered spectroscopy (DOSY) and nuclear Overhauser effect spectroscopy (NOESY), complemented by cryo-TEM were employed to address this. Efavirenz as model drug, the triblock copolymers Pluronic F-127 (PF127) and poly(2-oxazolin) based pMeOx-b-pPrOzi-b-pMeOx (pOx/pOzi) and their respective formulations were studied in simulated fed-state intestinal fluid (FeSSIF). For the individual polymers, the bile interfering nature of PF127 was confirmed and pure pOx/pOzi was newly classified as non-interfering. A different and more complex behaviour was observed if EFV was involved. The formulations showed multi-facetted concentration and composition dependent aggregation. This demonstrates that separate evaluation of polymers or drugs in biorelevant media is not sufficient and their mixtures need to be carefully studied.<br>

The Total Solubility of the Co-Solubilized PAHs with Similar Structures Indicated by NMR Chemical Shift

The synergism/inhibition level, solubilization sites and the total solubility (St) of co-solubilization systems of phenanthrene, anthracene and pyrene in Tween 80 and sodium dodecyl sulfate (SDS) are studied by 1H-NMR, 2D nuclear overhauser effect spectroscopy (NOESY) and rotating frame overhauser effect spectroscopy (ROESY). In Tween 80, inhibition for phenanthrene, anthracene and pyrene is observed in most binary and ternary systems. However, in SDS, synergism is predominant. After analysis, we find that different synergism or inhibition situation between Tween 80 and SDS is related to the different types of surfactants used and the resulting different co-solubilization mechanisms. In addition, we also find that three polycyclic aromatic hydrocarbons.

Investigation of the structure of regulatory proteins interacting with glycosaminoglycans by combining NMR spectroscopy and molecular modeling – the beginning of a wonderful friendship

The interaction of regulatory proteins with extracellular matrix or cell surface-anchored glycosaminoglycans (GAGs) plays important roles in molecular recognition, wound healing, growth, inflammation and many other processes. In spite of their high biological relevance, protein-GAG complexes are significantly underrepresented in structural databases because standard tools for structure determination experience difficulties in studying these complexes. Co-crystallization with subsequent X-ray analysis is hampered by the high flexibility of GAGs. NMR spectroscopy experiences difficulties related to the periodic nature of the GAGs and the sparse proton network between protein and GAG with distances that typically exceed the detection limit of nuclear Overhauser enhancement spectroscopy. In contrast, computer modeling tools have advanced over the last years delivering specific protein-GAG docking approaches successfully complemented with molecular dynamics (MD)-based analysis. Especially the combination of NMR spectroscopy in solution providing sparse structural constraints with molecular docking and MD simulations represents a useful synergy of forces to describe the structure of protein-GAG complexes. Here we review recent methodological progress in this field and bring up examples where the combination of new NMR methods along with cutting-edge modeling has yielded detailed structural information on complexes of highly relevant cytokines with GAGs.