RESUMEN
One of the mechanisms accounting for the toxicity of amyloid peptides in diseases like Alzheimer's and Parkinson's is the formation of pores on the plasma membrane of neurons. Here, we perform unbiased all-atom simulations of the full membrane damaging pathway, which includes adsorption, aggregation, and perforation of the lipid bilayer accounting for pore-like structures. Simulations are performed using four peptides made with the same amino acids. Differences in the nonpolar-polar sequence pattern of these peptides prompt them to adsorb into the membrane with the extended conformations oriented either parallel [peptide labeled F1, Ac-(FKFE)2-NH2], perpendicular (F4, Ac-FFFFKKEE-NH2), or with an intermediate orientation (F2, Ac-FFKKFFEE-NH2, and F3, Ac-FFFKFEKE-NH2) in regard to the membrane surface. At the water-lipid interface, only F1 fully self-assembles into ß-sheets, and F2 peptides partially fold into an α-helical structure. The ß-sheets of F1 emerge as electrostatic interactions attract neighboring peptides to intermediate distances where nonpolar side chains can interact within the dry core of the bilayer. This complex interplay between electrostatic and nonpolar interactions is not observed for the other peptides. Although ß-sheets of F1 peptides are mostly parallel to the membrane, some of their edges penetrate deep inside the bilayer, dragging water molecules with them. This precedes pore formation, which starts with the flow of two water layers through the membrane that expand into a stable cylindrical pore delimited by polar faces of ß-sheets spanning both leaflets of the bilayer.
RESUMEN
RyR1 is an intracellular Ca2+ channel important in excitable cells such as neurons and muscle fibers. Ca2+ activates it at low concentrations and inhibits it at high concentrations. Mg2+ is the main physiological RyR1 inhibitor, an effect that is overridden upon activation. Despite the significance of Mg2+-mediated inhibition, the molecular-level mechanisms remain unclear. In this work we determined two cryo-EM structures of RyR1 with Mg2+ up to 2.8 Å resolution, identifying multiple Mg2+ binding sites. Mg2+ inhibits at the known Ca2+ activating site and we propose that the EF hand domain is an inhibitory divalent cation sensor. Both divalent cations bind to ATP within a crevice, contributing to the precise transmission of allosteric changes within the enormous channel protein. Notably, Mg2+ inhibits RyR1 by interacting with the gating helices as validated by molecular dynamics. This structural insight enhances our understanding of how Mg2+ inhibition is overcome during excitation.
Asunto(s)
Calcio , Microscopía por Crioelectrón , Magnesio , Canal Liberador de Calcio Receptor de Rianodina , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Canal Liberador de Calcio Receptor de Rianodina/química , Magnesio/metabolismo , Calcio/metabolismo , Sitios de Unión , Animales , Simulación de Dinámica Molecular , Adenosina Trifosfato/metabolismo , Humanos , ConejosRESUMEN
Rhipicephalus (Boophilus) microplus is tick parasite that affects the cattle industry worldwide. In R. (B.) microplus, acaricide resistance develops rapidly against many commercial acaricides. One of main resistance strategies is to enhance the metabolic detoxification mediated by R. (B.) microplus glutathione-S-transferase (RmGST). RmGST detoxifies acaricides by catalyzing the conjugation of glutathione to acaricides. Although structural and dynamic details of RmGST are expected to elucidate the biologic activity of this molecule, these data have not been available to date. Thus, Molecular Dynamics simulations were employed to study ligand-free RmGST at an atomic level. Like other m-class GSTs, the flexible m loop (m1) of RmGST was observed. M1 seems to shield the active sites from the bulk. A RmGST dimer is stabilized by the lock-and-key motif (F57 as "key") and hydrogen bonds of R82-E91 and R82-D98 at the dimer interface. Without substrates, conserved catalytic Y116 and N209 can interact with V112, G210 (for Y116) and F215 (for N209). Overall, most residues involving in RmGST function and stability are similar to other m-class GSTs. This implies similar structural stability and catalytic activity of RmGST to other GSTs. An insight obtained here will be useful for management of acaricide resistance and tick control.Communicated by Ramaswamy H. Sarma.
RESUMEN
Rhipicephalus (Boophilus) microplus is one of the most widespread ticks causing a massive loss to livestock production. The long-term use of acaracides rapidly develops acaracide resistance. In R. microplus, enhancing the metabolic activity of glutathione S-transferase (RmGST) is one of the mechanisms underlying acaracide resistance. RmGST catalyzes the conjugation of glutathione (GSH) to insecticides causing an easy-to-excrete conjugate. The active RmGST dimer contains two active sites (hydrophobic co-substrate binding site (H-site) and GSH binding site (G-site)) in each monomer. To preserve the insecticide efficacy, s-hexyl glutathione (GTX), a GST inhibitor, has been used as a synergist. To date, no molecular information on the RmGST-GSH/GTX complex is available. The insight is important for developing a novel RmGST inhibitor. Therefore, in this work, molecular dynamics simulations (MD) were performed to explore the binding of GTX and GSH to RmGST. GSH binds tighter and sits rigidly inside the G-site, while flexible GTX occupies both active sites. In GSH, the backbone mainly interacts with W8, R43, W46, K50, N59, L60, Q72, and S73, while its thiol group directs to Y7. In contrast, the aliphatic hexyl of GTX protrudes into the H-site and allows a flexible peptide core to form various interactions. Such high GTX flexibility and the protrusion of its hexyl moiety to the H-site suggest the dual role of GTX in preventing the conjugation reaction and the binding of acaracide. This insight can provide a better understanding of an important insecticide-resistance mechanism, which may in turn facilitate the development of novel approaches to tick control.
Asunto(s)
Acaricidas , Insecticidas , Rhipicephalus , Animales , Rhipicephalus/metabolismo , Glutatión Transferasa/metabolismo , Insecticidas/farmacología , Resistencia a los Insecticidas , Acaricidas/farmacología , Glutatión/metabolismoRESUMEN
Membrane scaffold proteins (MSP) nanodiscs have been extensively used in structural study of membrane proteins. In cryo-EM, an incorporation of target proteins into nanodiscs is conducted under a rapid change from cryogenic to ambient temperatures. We present a coarse-grained molecular dynamics (CGMD) study for investigating an effect of temperature on the structural organization of DPPC-nanodisc and POPC-nanodisc. A non-monotonic response of physical quantities (i.e. the lipid order parameter, nanodisc flatness, structural change, solvation property, radius of gyration) with increase in temperature (T = 200-350 K) is found to be associated with the gel-ripple-liquid crystalline phase change within nanodiscs. The reorganization of lipids upon temperature variation induced conformational changes of MSP to minimize hydrophobic exposure of the lipid membrane to an aqueous environment. Structural response to temperature is different to a certain extent between the saturated DPPC and unsaturated POPC.
Asunto(s)
1,2-Dipalmitoilfosfatidilcolina/química , Simulación de Dinámica Molecular , Nanoestructuras/química , Fosfatidilcolinas/química , Temperatura , Interacciones Hidrofóbicas e HidrofílicasRESUMEN
Structural response of a AQP1 is examined by a coarse-grained model with a phenomenological interaction potential with a knowledge-based residue-residue interaction (derived from an ensemble of protein structures in PDB). The thermal response of the protein chain exhibits an unexpected characteristics in its native phase where the radius of gyration of the protein decreases on raising the temperature. The radius of gyration of AQP1 increases on increasing the temperature before saturating to a random-coil morphology in denatured phase at high temperatures. Three regions of persistent globularization are identified, toward the end segments 1M-25V and 250V-269K and a narrow region in the middle 155A-163D along the backbone. Varying the temperature leads to a systematic redistribution of self-organizing residues with globular and fibrous morphologies with an effective dimension D~2 (random coil) at high temperature and D~3 (globular conformation) in native phase. A preliminary analysis is also presented on the effect of a crowded membrane environment on the protein structure by incorporating effective solute constituents. Conformation of the protein is found to be pinned by selective binding of solute to specific targets; the matrix directed structure differs considerably from that of a protein in a generic solvent. The structure of AQP1 can be controlled by temperature and constitutive elements of the underlying matrix.
Asunto(s)
Acuaporina 1 , Conformación Proteica , Pliegue de Proteína , Acuaporina 1/química , Modelos Moleculares , Solventes , TemperaturaRESUMEN
A new series of furofuran lignans containing catechol moiety were prepared from the reactions between lignans and a variety of phenolics. All 22 products obtained were evaluated against three different α-glucosidases (maltase, sucrase and Baker's yeast glucosidase) and DPPH radical. Of furofuran lignans evaluated, ß-14, having two catechol moieties and one acetoxy group, was the most potent inhibitor against Baker's yeast, maltase, and sucrase with IC50 values of 5.3, 25.7, and 12.9⯵M, respectively. Of interest, its inhibitory potency toward Baker's yeast was 28 times greater than standard drug, acarbose and its DPPH radical scavenging (SC50 11.2⯵M) was 130 times higher than commercial antioxidant BHT. Subsequent investigation on mechanism underlying the inhibitory effect of ß-14 revealed that it blocked Baker's yeast and sucrase functions by mixed-type inhibition while it exerted non-competitive inhibition toward maltase. Molecular dynamics simulation of the most potent furofuran lignans (4, α-8b, α-14, and ß-14) with the homology rat intestinal maltase at the binding site revealed that the hydrogen bond interactions from catechol, acetoxy, and quinone moieties of furofuran lignans were the key interaction to bind tightly to α-glucosidase. The results indicated that ß-14 possessed promising antidiabetic activity through simultaneously inhibiting α-glucosidases and free radicals.