Biophysics and physicobiology最新文献

Deposition and accumulation of amyloid fibrils is a hallmark of a group of diseases called amyloidosis and neurodegenerative disorders. Although polypeptides potentially have a fibril-forming propensity, native proteins have evolved into proper functional conformations to avoid aggregation and fibril formation. Understanding the mechanism for regulation of fibril formation of native proteins provides clues for the rational design of molecules for inhibiting fibril formation. Although fibril formation is a complex multistep reaction, experimentally obtained fibril formation curves can be fitted with the Finke-Watzky (F-W) two-step model for homogeneous nucleation followed by autocatalytic fibril growth. The resultant F-W rate constants for nucleation and fibril formation provide information on the chemical kinetics of fibril formation. Using the F-W two-step model analysis, we investigated the physicochemical mechanisms of fibril formation of a Parkinson's disease protein α-synuclein (αS) and a systemic amyloidosis protein apolipoprotein A-I (apoA-I). The results indicate that the C-terminal region of αS enthalpically and entropically suppresses nucleation through the intramolecular interaction with the N-terminal region and the intermolecular interaction with existing fibrils. In contrast, the nucleation of the N-terminal fragment of apoA-I is entropically driven likely due to dehydration of large hydrophobic segments in the molecule. Based on our recent findings, we discuss the similarity and difference of the fibril formation mechanisms of αS and the N-terminal fragment of apoA-I from the physicochemical viewpoints.

Cell migration plays an important role in the development and maintenance of multicellular organisms. Factors that induce cell migration and mechanisms controlling their expression are important for determining the mechanisms of factor-induced cell migration. Despite progress in the study of factor-induced cytotaxis, including chemotaxis and haptotaxis, precise control of the direction of cell migration over a wide area has not yet been achieved. Success in this area would update the cell migration assays, superior cell separation technologies, and artificial organs with high biocompatibility. The present study therefore sought to control the direction of cell migration over a wide area by adjusting the three-dimensional shape of the cell scaffold. The direction of cell migration was influenced by the shape of the cell scaffold, thereby optimizing cell adhesion and protrusion. Anisotropic arrangement of these three-dimensional shapes into a periodic structure induced unidirectional cell migration. Three factors were required for unidirectional cell migration: 1) the sizes of the anisotropic periodic structures had to be equal to or lower than the size of the spreading cells, 2) cell migration was restricted to a runway approximately the width of the cell, and 3) cells had to be prone to extension of long protrusions in one direction. Because the first two factors had been identified previously in studies of cell migration in one direction using two-dimensional shaped patterns, these three factors are likely important for the mechanism by which cell scaffold shapes regulate cell migration.

How do the living systems emerge from non-living molecular assemblies? What physical and chemical principles supported the process? To address these questions, a promising strategy is to artificially reconstruct living cells in a bottom-up way. Recently, the authors developed the "synthetic minimal cell" system showing recursive growth and division cycles, where the concepts of information molecules, metabolic pathways, and cell reproduction were artificially and concisely redesigned with the vesicle-based system. We intentionally avoided using the sophisticated molecular machinery of the biological cells and tried to redesign the cells in the simplest forms. This review focuses on the similarities and differences between the biological cells and our synthetic minimal cell concerning each concept of cells. Such comparisons between natural and artificial cells will provide insights on how the molecules should be assembled to create living systems to the wide readers in the field of synthetic biology, artificial cells, and protocells research. This review article is an extended version of the Japanese article "Growth and division of vesicles coupled with information molecules," published in SEIBUTSU-BUTSURI vol. 61, p. 378-381 (2021).

KaiC is a multifunctional enzyme functioning as the core of the circadian clock system in cyanobacteria: its N-terminal domain has adenosine triphosphatase (ATPase) activity, and its C-terminal domain has autokinase and autophosphatase activities targeting own S431 and T432. The coordination of these multiple biochemical activities is the molecular basis for robust circadian rhythmicity. Therefore, much effort has been devoted to elucidating the cooperative relationship between the two domains. However, structural and functional relationships between the two domains remain unclear especially with respect to the dawn phase, at which KaiC relieves its nocturnal history through autodephosphorylation. In this study, we attempted to design a double mutation of S431 and T432 that can capture KaiC as a fully dephosphorylated form with minimal impacts on its structure and function, and investigated the cooperative relationship between the two domains in the night to morning phases from many perspectives. The results revealed that both domains cooperate at the dawn phase through salt bridges formed between the domains, thereby non-locally co-activating two events, ATPase de-inhibition and S431 dephosphorylation. Our further analysis using existing crystal structures of KaiC suggests that the states of both domains are not always in one-to-one correspondence at every phase of the circadian cycle, and their coupling is affected by the interactions with KaiA or adjacent subunits within a KaiC hexamer.

A small and flexible molecule, ribocil A (non-binder) or B (binder), binds to the deep pocket of the aptamer domain of the FMN riboswitch, which is an RNA molecule. This binding was studied by mD-VcMD, which is a generalized-ensemble simulation method. Ribocil A and B are structurally similar because they are optical isomers to each other. In the initial conformation of simulation, the ligands and the aptamer were completely dissociated in explicit solvent. The aptamer-ribocil B binding was stronger than the aptamer-ribocil A binding, which agrees with experiments. The computed free-energy landscape for the aptamer-ribocil B binding was funnel-like, whereas that for the aptamer-ribocil A binding was rugged. When passing through the gate (named "front gate") of the binding pocket, each ligand interacted with bases of the riboswitch by non-native π-π stackings, and the stackings restrained the ligand's orientation to be advantageous to reach the binding site smoothly. When the ligands reached the binding site in the pocket, the non-native stackings were replaced by the native stackings. The ligand's orientation restriction is discussed referring to a selection mechanism reported in an earlier work on a drug-GPCR interaction. The present simulation showed another pathway leading the ligands to the binding site. The gate ("rear gate") for this pathway was located completely opposite to the front gate on the aptamer's surface. However, the approach from the rear gate required overcoming a free-energy barrier regarding ligand's rotation before reaching the binding site.

The consistency principle represents a physicochemical condition requisite for ideal protein folding. It assumes that any pair of amino acid residues in partially folded structures has an attractive short-range interaction only if the two residues are in contact within the native structure. The residue-specific equilibrium constant, K, and the residue-specific rate constant, k (forward and backward), can be determined by NMR and hydrogen-deuterium exchange studies. Linear free energy relationships (LFER) in the rate-equilibrium free energy relationship (REFER) plots (i.e., log k vs. log K) are widely seen in protein-related phenomena, but our REFER plot differs from them in that the data points are derived from one polypeptide chain under a single condition. Here, we examined the theoretical basis of the residue-based LFER. First, we derived a basic equation, ρ_ij=½(φ_i+φ_j), from the consistency principle, where ρ_ij is the slope of the line segment that connects residues i and j in the REFER plot, and φ_i and φ_j are the local fractions of the native state in the transient state ensemble (TSE). Next, we showed that the general solution is the alignment of the (log K, log k) data points on a parabolic curve in the REFER plot. Importantly, unlike LFER, the quadratic free energy relationship (QFER) is compatible with the heterogeneous formation of local structures in the TSE. Residue-based LFER/QFER provides a unique insight into the TSE: A foldable polypeptide chain consists of several folding units, which are consistently coupled to undergo smooth structural changes.

Aggregates of amyloid-β (Aβ) peptides are thought to cause Alzheimer's disease. Polyphenolic compounds are known to inhibit Aβ aggregation. We applied replica permutation with solute tempering (RPST) to the system of Aβ fragments, Aβ(16-22), and polyphenols to elucidate the mechanism of inhibition of Aβ aggregation. The RPST molecular dynamics simulations were performed for two polyphenols, myricetin (MYC) and rosmarinic acid (ROA). Two Aβ fragments were distant, and the number of residues forming the intermolecular β-sheet was reduced in the presence of MYC and ROA compared with that in the absence of polyphenols. MYC was found to interact with glutamic acid and phenylalanine of Aβ fragments. These interactions induce helix structure formation of Aβ fragments, making it difficult to form β-sheet. ROA interacted with glutamic acid and lysine, which reduced the hydrophilic interaction between Aβ fragments. These results indicate that these polyphenols inhibit the aggregation of Aβ fragments with different mechanisms.