Helix stabilization

Despite the importance of proline conformational equilibria ( trans versus cis amide and exo versus endo ring pucker) on protein structure and function, there is a lack of convenient ways to probe proline conformation. 4,4-Difluoroproline (Dfp) was identified to be a sensitive 19 F NMR-based probe of proline conformational biases and cis-trans isomerism. Within model compounds and disordered peptides, the diastereotopic fluorines of Dfp exhibit similar chemical shifts (ΔδFF = 0-3 ppm) when a trans X-Dfp amide bond is present...

HIV-1 Vpr is a multifunctional accessory protein consisting of 96 amino acids that play a critical role in viral pathogenesis. Among its diverse range of activities, Vpr can create a cation-selective ion channel within the plasma membrane. However, the oligomeric state of this channel has not yet been elucidated. In this study, we investigated the conformational dynamics of Vpr helices to model the ion channel topology. First, we employed a series of multiscale simulations to investigate the specific structure of monomeric Vpr in a membrane model...

Giant lipid vesicles composed of a lipid bilayer form complex membrane structures and enzyme network reactions that can be used to construct well-defined artificial cell models based on microfluidic technologies and synthetic biology. As a different approach to cell-mimicking systems, we formed an asymmetric lipid-amphiphilic protein (oleosin) vesicle containing a lipid and an oleosin monolayer in the outer and inner leaflets, respectively. These asymmetric vesicles enabled the reconstitution and function of β-barrel types of membrane proteins (OmpG) and the fission of vesicles stimulated by lysophospholipids...

Glycine rich polyproline II helix assemblies are an emerging class of natural domains found in several proteins with different functions and diverse origins. The distinct properties of these domains relative to those composed of α-helices and β-sheets could make glycine-rich polyproline II helix assemblies a useful building block for protein design. Whereas the high population of polyproline II conformers in disordered state ensembles could facilitate glycine-rich polyproline II helix folding, the architectonic bases of these structures are not well known...

Choline is a vital nutrient and a precursor for the biosynthesis of essential metabolites, including acetylcholine (ACh), that play a central role in fetal development, especially in the brain. In cholinergic neurons, the high-affinity choline transporter (CHT1) provides an extraordinarily efficient reuptake mechanism to reutilize choline derived from intrasynaptical ACh hydrolysis and maintain ACh synthesis in the presynapse. Here, we determined structures of human CHT1 in three discrete states: the outward-facing state bound with the competitive inhibitor hemicholinium-3 (HC-3); the inward-facing occluded state bound with the substrate choline; and the inward-facing apo open state...

UNLABELLED: Fungal resistance to commonly used medicines is a growing public health threat, and there is a dire need to develop new classes of antifungals. We previously described a peptide produced by Enterococcus faecalis , EntV, that restricts Candida albicans to a benign form rather than having direct fungicidal activity. Moreover, we showed that one 12-amino acid (aa) alpha helix of this peptide retained full activity, with partial activity down to the 10aa alpha helix. Using these peptides as a starting point, the current investigation sought to identify the critical features necessary for antifungal activity and to screen for new variants with enhanced activity using both biofilm and C...

The viral genome of SARS-CoV-2 is packaged by the nucleocapsid (N-)protein into ribonucleoprotein particles (RNPs), 38 ± 10 of which are contained in each virion. Their architecture has remained unclear due to the pleomorphism of RNPs, the high flexibility of N-protein intrinsically disordered regions, and highly multivalent interactions between viral RNA and N-protein binding sites in both N-terminal (NTD) and C-terminal domain (CTD). Here we explore critical interaction motifs of RNPs by applying a combination of biophysical techniques to ancestral and mutant proteins binding different nucleic acids in an in vitro assay for RNP formation, and by examining nucleocapsid protein variants in a viral assembly assay...

We describe molecular-level functional changes in the α4β2 nicotinic acetylcholine receptor (nAChR) by a leucine residue insertion in the M2 transmembrane domain of the α4 subunit associated with sleep-related hyperkinetic epilepsy. Measurements of agonist-elicited single channel currents reveal the primary effect is to stabilize the open channel state while the secondary effect is to promote reopening of the channel. These dual effects prolong the durations of bursts of channel openings equally for the two major stoichiometric forms of the receptor, (α4)2 (β2)3 and (α4)3 (β2)2 , indicating the functional impact is independent of mutant copy number per receptor...

In this work, the effects of low-frequency alternating magnetic fields (LF-AMF) on the physicochemical, conformational, and functional characteristics of myofibrillar protein (MP) after iterative freeze-thaw (FT) cycles were explored. With the increasing LF-AMF treatment time, the solubility, active sulfhydryl groups, surface hydrophobicity, emulsifiability, and emulsion stability of MP after five FT cycles evidently elevated and then declined, and the peak value was obtained at 3 h. Conversely, the moderate LF-AMF treatment time can significantly reduce the average particle size, carbonyl content, and endogenous fluorescence intensity of MP...

The obtained seeds from fruit processing are considered by-products containing proteins that could be utilized as ingredients in food manufacturing. However, in the specific case of soursop seeds, their usage for the preparation of protein isolates is limited. In this investigation a protein isolate from soursop seeds (SSPI) was obtained by alkaline extraction and isoelectric precipitation methods. The SSPI was sonicated at 200, 400 and 600 W during 15 and 30 min and its effect on the physicochemical, functional, biochemical, and structural properties was evaluated...

The rice glutelin (RG), the separated retentate (RGFs) and filtrate (FGFs) fractions from total glutelin fibrils (TGFs) at pH 3.5 were used as carrier for curcumin in this test. The solubility and antioxidant activities of curcumin were improved after binding with protein and fibrils. Compared to other complexes, the RGFs-curcumin complex exhibited a highest curcumin solubility (48.05%) and a superior sustained release property, probably owing to the stable hydrogen bond between the surface groups of fibrils and hydroxyl groups of polyphenols...

Optically active poly(naphthalene-1,4-diyl) was prepared through helix-sense-selective polymerization of the corresponding monomers and also through circularly polarized light (CPL) irradiation, resulting in distinctive circular dichroism (CD) spectral patterns. Chirality of the helix-sense-selective polymerization -based polymer is ascribed to preferred-handed helicity while that of the CPL-based polymer to a non-helical, chiral conformation ('biased-dihedral conformation') with preferred-handedness which was stable only in the solid state...

We study numerically nonlinear dynamics of several types of molecular systems composed of hydrogen-bonded chains placed inside carbon nanotubes with open edges. We demonstrate that carbon nanotubes provide a stabilization mechanism for quasi-one-dimensional molecular chains via the formation of their secondary structures. In particular, a polypeptide chain (Gly)N placed inside a carbon nanotube can form a stable helical chain (310-, α-, π-, and β-helix) with parallel chains of hydrogen-bonded peptide groups...

30S subunits become inactive upon exposure to low Mg2+ concentration, due to a reversible conformational change that entails nucleotides (nt) in the neck helix (h28) and 3' tail of 16S rRNA. This active-to-inactive transition involves partial unwinding of h28 and re-pairing of nt 921-923 with nt 1532-1534, which requires flipping of the 3' tail by ~180 degrees. Growing evidence suggests that immature 30S particles adopt the inactive conformation in the cell, and transition to the active state occurs at a late stage of maturation...

BACKGROUND: This study explored the denaturation of 11S globulin, a protein known for its diverse functionalities in soy protein applications, at pH 3.0 and pH 10.0, subsequently followed by a gradual return to pH 7.0 to facilitate renaturation. It investigated the structural and functional changes during renaturation induced by pH, unveiling the stabilization mechanism of 11S globulin. RESULTS: The findings revealed that during pH adjustment to neutral, the denatured soybean 11S globulin-resulting from alkaline (pH 10...

Mucus is a complex biological hydrogel that acts as a barrier for almost everything entering or exiting the body. It is therefore of emerging interest for biomedical and pharmaceutical applications. Besides water, the most abundant components are the large and densely glycosylated mucins, glycoproteins of up to 20 MDa and carbohydrate content of up to 80 wt%. Here, we designed and explored a library of glycosylated peptides to deconstruct the complexity of mucus. Using the well-characterized hFF03 coiled-coil system as a hydrogel-forming peptide scaffold, we systematically probed the contribution of single glycans to the secondary structure as well as the formation and viscoelastic properties of the resulting hydrogels...

The B domain of protein A (BdpA), a small three-helix bundle, folds on a time scale of a few microseconds with heterogeneous native and unfolded states. It is widely used as a model for understanding protein folding mechanisms. In this work, we use structure-based models (SBMs) and atomistic simulations to comprehensively investigate how BdpA folding is associated with the formation of its secondary structure. The energy landscape visualization method (ELViM) was used to characterize the pathways that connect the folded and unfolded states of BdpA as well as the sets of structures displaying specific ellipticity patterns...

The field of protein design has made remarkable progress over the past decade. Historically, the low reliability of purely structure-based design methods limited their application, but recent strategies that combine structure-based and sequence-based calculations, as well as machine learning tools, have dramatically improved protein engineering and design. In this Review, we discuss how these methods have enabled the design of increasingly complex structures and therapeutically relevant activities. Additionally, protein optimization methods have improved the stability and activity of complex eukaryotic proteins...

Amorphous solid dispersions (ASDs) using proteins as carriers have emerged as a promising strategy for stabilizing amorphous drug molecules. Proteins possess diverse three-dimensional structures that significantly influence their own properties and may also impact the properties of ASDs. We prepared β-lactoglobulin (BLG) with different contents of β-sheet and α-helical secondary structures by initially dissolving BLG in different mixed solvents, containing different ratios of water, methanol/ethanol, and acetic acid, followed by spray drying of the solutions...

The human primosome, a four-subunit complex of primase and DNA polymerase alpha (Polα), initiates DNA synthesis on both chromosome strands by generating chimeric RNA-DNA primers for loading DNA polymerases delta and epsilon (Polε). Replication protein A (RPA) tightly binds to single-stranded DNA strands, protecting them from nucleolytic digestion and unauthorized transactions. We report here that RPA plays a critical role for the human primosome during DNA synthesis across inverted repeats prone to hairpin formation...