A molten globule ensemble primes Arf1-GDP for the nucleotide switch.

The adenosine di-phosphate (ADP) ribosylation factor (Arf) small guanosine tri-phosphate (GTP)ases function as molecular switches to activate signaling cascades that control membrane organization in eukaryotic cells. In Arf1, the GDP/GTP switch does not occur spontaneously but requires guanine nucleotide exchange factors (GEFs) and membranes. Exchange involves massive conformational changes, including disruption of the core β-sheet.

Host Defense Peptide Piscidin and Yeast-Derived Glycolipid Exhibit Synergistic Antimicrobial Action through Concerted Interactions with Membranes.

Developing new antimicrobials as alternatives to conventional antibiotics has become an urgent race to eradicate drug-resistant bacteria and to save human lives. Conventionally, antimicrobial molecules are studied independently even though they can be cosecreted . In this research, we investigate two classes of naturally derived antimicrobials: sophorolipid (SL) esters as modified yeast-derived glycolipid biosurfactants that feature high biocompatibility and low production cost; piscidins, which are host defense peptides (HDPs) from fish.

Pressure pushes tRNA into excited conformational states.

Conformational dynamics play essential roles in RNA function. However, detailed structural characterization of excited states of RNA remains challenging. Here, we apply high hydrostatic pressure (HP) to populate excited conformational states of tRNA, and structurally characterize them using a combination of HP 2D-NMR, HP-SAXS (HP-small-angle X-ray scattering), and computational modeling.

High Pressure CPMG and CEST Reveal That Cavity Position Dictates Distinct Dynamic Disorder in the PP32 Repeat Protein.

Given the central role of conformational dynamics in protein function, it is essential to characterize the time scales and structures associated with these transitions. High pressure (HP) perturbation favors transitions to excited states because they typically occupy a smaller molar volume, thus facilitating characterization of conformational dynamics. Repeat proteins, with their straightforward architecture, provide good models for probing the sequence dependence of protein conformational dynamics.

Probing IgG1 F-Multimodal Nanoparticle Interactions: A Combined Nuclear Magnetic Resonance and Molecular Dynamics Simulations Approach.

In this study, NMR and molecular dynamics simulations were employed to study IgG1 F binding to multimodal surfaces. Gold nanoparticles functionalized with two multimodal cation-exchange ligands (Capto and Nuvia) were synthesized and employed to carry out solution-phase NMR experiments with the F. Experiments with perdeuterated N-labeled F and the multimodal surfaces revealed micromolar residue-level binding affinities as compared to millimolar binding affinities with these ligands in free solution, likely due to cooperativity and avidity effects.

Identification of preferred multimodal ligand-binding regions on IgG1 F using nuclear magnetic resonance and molecular dynamics simulations.

In this study, the binding of multimodal chromatographic ligands to the IgG1 F domain were studied using nuclear magnetic resonance and molecular dynamics simulations. Nuclear magnetic resonance experiments carried out with chromatographic ligands and a perdeuterated N-labeled F domain indicated that while single-mode ion exchange ligands interacted very weakly throughout the F surface, multimodal ligands containing negatively charged and aromatic moieties interacted with specific clusters of residues with relatively high affinity, forming distinct binding regions on the F . The multimodal ligand-binding sites on the F were concentrated in the hinge region and near the interface of the C 2 and C 3 domains.

Sequence-independent recognition of the amyloid structural motif by GFP protein family.

Cnidarian fluorescent protein (FP) derivatives such as GFP, mCherry, and mEOS2 have been widely used to monitor gene expression and protein localization through biological imaging because they are considered functionally inert. We demonstrate that FPs specifically bind amyloid fibrils formed from many natural peptides and proteins. FPs do not bind other nonamyloid fibrillar structures such as microtubules or actin filaments and do not bind to amorphous aggregates.

Elucidating the unusual reaction kinetics of D-glucuronyl C5-epimerase.

The chemoenzymatic synthesis of heparin, through a multienzyme process, represents a critical challenge in providing a safe and effective substitute for this animal-sourced anticoagulant drug. D-glucuronyl C5-epimerase (C5-epi) is an enzyme acting on a heparin precursor, N-sulfoheparosan, catalyzing the reversible epimerization of D-glucuronic acid (GlcA) to L-iduronic acid (IdoA). The absence of reliable assays for C5-epi has limited elucidation of the enzymatic reaction and kinetic mechanisms.

Pressure-Temperature Analysis of the Stability of the CTL9 Domain Reveals Hidden Intermediates.

The observation of two-state unfolding for many small single-domain proteins by denaturants has led to speculation that protein sequences may have evolved to limit the population of partially folded states that could be detrimental to fitness. How such strong cooperativity arises from a multitude of individual interactions is not well understood. Here, we investigate the stability and folding cooperativity of the C-terminal domain of the ribosomal protein L9 in the pressure-temperature plane using site-specific NMR.

The consequences of cavity creation on the folding landscape of a repeat protein depend upon context.

The effect of introducing internal cavities on protein native structure and global stability has been well documented, but the consequences of these packing defects on folding free-energy landscapes have received less attention. We investigated the effects of cavity creation on the folding landscape of the leucine-rich repeat protein pp32 by high-pressure (HP) and urea-dependent NMR and high-pressure small-angle X-ray scattering (HPSAXS). Despite a modest global energetic perturbation, cavity creation in the N-terminal capping motif (N-cap) resulted in very strong deviation from two-state unfolding behavior.

High-Pressure NMR and SAXS Reveals How Capping Modulates Folding Cooperativity of the pp32 Leucine-rich Repeat Protein.

Many repeat proteins contain capping motifs, which serve to shield the hydrophobic core from solvent and maintain structural integrity. While the role of capping motifs in enhancing the stability and structural integrity of repeat proteins is well documented, their contribution to folding cooperativity is not. Here we examined the role of capping motifs in defining the folding cooperativity of the leucine-rich repeat protein, pp32, by monitoring the pressure- and urea-induced unfolding of an N-terminal capping motif (N-cap) deletion mutant, pp32-∆N-cap, and a C-terminal capping motif destabilization mutant pp32-Y131F/D146L, using residue-specific NMR and small-angle X-ray scattering.

Zonal variation of MRI-measurable parameters classifies cartilage degradation.

Osteoarthritis (OA) is a degenerative joint disease resulting in the deterioration of articular cartilage, a tissue with minimal ability to self-repair. Early diagnosis of OA with non-invasive imaging techniques such as magnetic resonance imaging (MRI) could provide an opportunity to intervene and slow or reverse this degeneration process. This study examines the classification of degradation states using MRI measurements.

Nuclease activity gives an edge to host-defense peptide piscidin 3 over piscidin 1, rendering it more effective against persisters and biofilms.

Host-defense peptides (HDPs) feature evolution-tested potency against life-threatening pathogens. While piscidin 1 (p1) and piscidin 3 (p3) are homologous and potent fish HDPs, only p1 is strongly membranolytic. Here, we hypothesize that another mechanism imparts p3 strong potency.

Temperature and pressure limits of guanosine monophosphate self-assemblies.

Guanosine monophosphate, among the nucleotides, has the unique property to self-associate and form nanoscale cylinders consisting of hydrogen-bonded G-quartet disks, which are stacked on top of one another. Such self-assemblies describe not only the basic structural motif of G-quadruplexes formed by, e.g.

NMR and Computation Reveal a Pressure-Sensitive Folded Conformation of Trp-Cage.

Beyond defining the structure and stability of folded states of proteins, primary amino acid sequences determine all of the features of their conformational landscapes. Characterizing how sequence modulates the population of protein excited states or folding pathways requires atomic level detailed structural and energetic information. Such insight is essential for improving protein design strategies, as well as for interpreting protein evolution.

High Pressure ZZ-Exchange NMR Reveals Key Features of Protein Folding Transition States.

Understanding protein folding mechanisms and their sequence dependence requires the determination of residue-specific apparent kinetic rate constants for the folding and unfolding reactions. Conventional two-dimensional NMR, such as HSQC experiments, can provide residue-specific information for proteins. However, folding is generally too fast for such experiments.

Poly(ω-pentadecalactone)--poly(l-lactide) Block Copolymers via Organic-Catalyzed Ring Opening Polymerization and Potential Applications.

Poly(pentadecalactone)--poly(l-lactide) (PPDL--PLLA) diblock copolymers were prepared via the organic catalyzed ring-opening polymerization (ROP) of l-lactide (l-LA) from PPDL macroinitiators using either 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) or 1,5,7-triazabicyclo[4.

Polylactide/Poly(ω-hydroxytetradecanoic acid) Reactive Blending: A Green Renewable Approach to Improving Polylactide Properties.

A green manufacturing technique, reactive extrusion (REx), was employed to improve the mechanical properties of polylactide (PLA). To achieve this goal, a fully biosourced PLA based polymer blend was conceived by incorporating small quantities of poly(ω-hydroxytetradecanoic acid) (PC14). PLA/PC14 blends were compatibilized by transesterification reactions promoted by 200 ppm titanium tetrabutoxide (Ti(OBu)4) during REx.

Rapid and accurate determination of the lignin content of lignocellulosic biomass by solid-state NMR.

Biofuels and biomaterials, produced from lignocellulosic feedstock, require facile access to cellulose and hemicellulose to be competitive with petroleum processing and sugar-based fermentation. Physical-chemical barriers resulting from lignin complicates the hydrolysis biomass into fermentable sugars. Thus, the amount of lignin within a substrate is critical in determining biomass processing.

Structural and thermodynamic characterization of the recognition of the S100-binding peptides TRTK12 and p53 by calmodulin.

Calmodulin (CaM) is a multifunctional messenger protein that activates a wide variety of signaling pathways in eukaryotic cells in a calcium-dependent manner. CaM has been proposed to be functionally distinct from the S100 proteins, a related family of eukaryotic calcium-binding proteins. Previously, it was demonstrated that peptides derived from the actin-capping protein, TRTK12, and the tumor-suppressor protein, p53, interact with multiple members of the S100 proteins.

NMR investigation of the role of osteocalcin and osteopontin at the organic-inorganic interface in bone.

Mechanical resilience of bone tissue decreases with age. The ability to comprehensively probe and understand bone properties could help alleviate this problem. One important aspect of bone quality that has recently been made evident is the presence of dilatational bands formed by osteocalcin (OC) and osteopontin (OPN), which contribute to fracture toughness.

Identifying specific protein residues that guide surface interactions and orientation on silica nanoparticles.

We identify specific acylphosphatase (AcP) residues that interact with silica nanoparticles (SNPs) using a combined NMR spectroscopy and proteomics-mass spectrometry approach. AcP associated with 4- and 15-nm diameter SNPs through a common and specific interaction surface formed by amino acids from the two α-helices of the protein. Greater retention of native protein structure was obtained on 4-nm SNPs than on 15-nm particles, presumably due to greater surface curvature-induced protein stabilization with the smaller SNPs.

Bacterial expression and purification of the amyloidogenic peptide PAPf39 for multidimensional NMR spectroscopy.

PAPf39 is a 39 residue peptide fragment from human prostatic acidic phosphatase that forms amyloid fibrils in semen. These fibrils have been implicated in facilitating HIV transmission. To enable structural studies of PAPf39 by NMR spectroscopy, efficient methods allowing the production of milligram quantities of isotopically labeled peptide are essential.