Showing papers on "Protein filament published in 1996"

Torsional rigidity of single actin filaments and actin–actin bond breaking force under torsion measured directly by in vitro micromanipulation

Abstract: Knowledge of the elastic properties of actin filaments is crucial for considering its role in muscle contraction, cellular motile events, and formation of cell shape. The stiffness of actin filaments in the directions of stretching and bending has been determined. In this study, we have directly determined the torsional rigidity and breaking force of single actin filaments by measuring the rotational Brownian motion and tensile strength using optical tweezers and microneedles, respectively. Rotational angular fluctuations of filaments supplied the torsional rigidity as (8.0 ± 1.2) × 10−26 Nm2. This value is similar to that deduced from the longitudinal rigidity, assuming the actin filament to be a homogeneous rod. The breaking force of the actin–actin bond was measured while twisting a filament through various angles using microneedles. The breaking force decreased greatly under twist, e.g., from 600–320 pN when filaments were turned through 90°, independent of the rotational direction. Our results indicate that an actin filament exhibits comparable flexibility in the rotational and longitudinal directions, but breaks more easily under torsional load.

Alteration of myosin cross bridges by phosphorylation of myosin-binding protein C in cardiac muscle.

Abstract: In addition to the contractile proteins actin and myosin, contractile filaments of striated muscle contain other proteins that are important for regulating the structure and the interaction of the two force-generating proteins In the thin filaments, troponin and tropomyosin form a Ca-sensitive trigger that activates normal contraction when intracellular Ca is elevated In the thick filament, there are several myosin-binding proteins whose functions are unclear Among these is the myosin-binding protein C (MBP-C) The cardiac isoform contains four phosphorylation sites under the control of cAMP and calmodulin-regulated kinases, whereas the skeletal isoform contains only one such site, suggesting that phosphorylation in cardiac muscle has a specific regulatory function We isolated natural thick filaments from cardiac muscle and, using electron microscopy and optical diffraction, determined the effect of phosphorylation of MBP-C on cross bridges The thickness of the filaments that had been treated with protein kinase A was increased where cross bridges were present No change occurred in the central bare zone that is devoid of cross bridges The intensity of the reflections along the 43-nm layer line, which is primarily due to the helical array of cross bridges, was increased, and the distance of the first peak reflection from the meridian along the 43-nm layer line was decreased The results indicate that phosphorylation of MBP-C (i) extends the cross bridges from the backbone of the filament and (ii) increases their degree of order and/or alters their orientation These changes could alter rate constants for attachment to and detachment from the thin filament and thereby modify force production in activated cardiac muscle

Filament compliance and tension transients in muscle.

Abstract: Huxley and Simmons (1971) proposed an explanation for some striking features in the transient tension response of a tetanized muscle fibre to sudden stretch or release. More recent work has shown that the stiffness of cross-bridges in skeletal muscle is an order of magnitude greater than appeared at that time. On the simplified treatment used in that paper, this would cause serious disagreement with the experimental observations. It is shown here analytically and by computer simulations that these discrepancies disappear when account is taken of (a) the range of positions of myosin molecules relative to attachment sites on the thin filament and (b) the recently-discovered compliance in the thin filaments.

Implications of intermediate filament protein phosphorylation.

Abstract: Intermediate filament (IF) proteins, a large family of tissue specific proteins, undergo several posttranslational modifications, with phosphorylation being the most studied modification. IF protein phosphorylation is highly dynamic and involves the head and/or tail domains of these proteins, which are the domains that impart most of the structural heterogeneity and hence presumed tissue specific functions. Although the function of IF proteins remains poorly understood, several regulatory roles for IF protein phosphorylation have been identified or are emerging. Those roles include filament disassembly and reorganization, solubility, localization within specific cellular domains, association with other cytoplasmic or membrane associated proteins, protection against physiologic stress and mediation of tissue-specific functions. Understanding the mechanistic and functional aspects of IF protein phosphorylation is providing insights not only regarding the function of this modification, but also regarding the function of IF proteins.

The contraction of liquid filaments

Abstract: In this paper the evolution of a free liquid filament of arbitrary viscosity, contracting under the action of surface tension forces, is studied by numerical means. A finite- element discretization procedure is used to obtain approximate solutions to the Navier-Stokes equations. A Lagrangian approach is employed to deal with the large domain deformations which occur during the evolution of the filament. Typically we find that during the contraction a bulbous region forms at the end of the filament. The character of the evolution of the filament is found to be crucially dependent on the value of the Ohnesorge number Oh (a measure of viscous and surface tension forces). For large Ohnesorge numbers (Oh [Gt ] O(1)) it is found that the liquid filament remains stable during contraction, even when the initial length of the filament is much longer than the Rayleigh stability limit. The bulbous end becomes more localized with decreasing Ohnesorge number while at the same time a clear neck forms in front of the bulbous end. In addition we find that the region in which the pressure is minimum moves towards the neck. For sufficiently small Ohnesorge numbers (Oh [Lt ] O(0.01)) the filament becomes unstable with the radius of the neck decreasing and, eventually, the bulbous end breaking away from the filament.

Myosin filament structure in vertebrate smooth muscle.

Abstract: The in vivo structure of the myosin filaments in vertebrate smooth muscle is unknown. Evidence from purified smooth muscle myosin and from some studies of intact smooth muscle suggests that they may have a nonhelical, side-polar arrangement of crossbridges. However, the bipolar, helical structure characteristic of myosin filaments in striated muscle has not been disproved for smooth muscle. We have used EM to investigate this question in a functionally diverse group of smooth muscles (from the vascular, gastrointestinal, reproductive, and visual systems) from mammalian, amphibian, and avian species. Intact muscle under physiological conditions, rapidly frozen and then freeze substituted, shows many myosin filaments with a square backbone in transverse profile. Transverse sections of fixed, chemically skinned muscles also show square backbones and, in addition, reveal projections (crossbridges) on only two opposite sides of the square. Filaments gently isolated from skinned smooth muscles and observed by negative staining show crossbridges with a 14.5-nm repeat projecting in opposite directions on opposite sides of the filament. Such filaments subjected to low ionic strength conditions show bare filament ends and an antiparallel arrangement of myosin tails along the length of the filament. All of these observations are consistent with a side-polar structure and argue against a bipolar, helical crossbridge arrangement. We conclude that myosin filaments in all smooth muscles, regardless of function, are likely to be side-polar. Such a structure could be an important factor in the ability of smooth muscles to contract by large amounts.

Structures and Mechanical Properties of Cellulose Filament Spun from Cellulose/Aqueous NaOH Solution System

Abstract: An attempt was made to disclose super-molecular structure and mechanical properties of new cellulose filament prepared from cellulose-aqueous alkali solution system. X-Ray crystallinity index χc(X) of the new cellulose filament was far higher than those of other commercial rayons, such as viscose including polynosic and cupro and was slightly lower than the organic spun rayon. The new cellulose filament showed the lowest degree of crystal orientation among these three kinds of cellulose fibers, because of its lowest draft and stretching ratio during the spinning process. In the new filament degree of the intramolecular hydrogen bondings (1−χam(C3)) estimated by 13C NMR method was highly developed. The mechanical loss tangent (tanδ) vs. temperature T curves of the new filament exhibited α relaxation (from higher temperature side; α2, αsh) attributed to micro-Brownian motion of the cellulose chains, two β (βa1, βa2), local mode of the chains, and γ, rotational mode of primary alcohol group at C6 position of pyranose rings, in the regions of T=250–100°C and −30∼−100°C, and near −100°C, respectively. Judging from the co relationships of these relaxation peak temperature Tmax to χc(X) and 1−χam(C3), we concluded that there exist two amorphous regions with well-developed intramolecular hydrogen bonds but less advanced intermolecular hydrogen bonds and vice-versa in the new filament. The results of thermally stimulated currency (TSC) measurements suggested that the hydrophobic interaction among cellulose chains is tightly formed in the region with the well-developed intramolecular hydrogen bonds. In this regard, χTSC peak named here was found to correspond to mechanical relaxation αsh. The tensile strength and elongation of the new filament were comparable to those of the regular viscose rayon. The new filament showed less swelling ratio and low fibrillation nature in water. Woven fabrics made from the filament gave some softness and high abrasion, compared with other commercial ones and were hard to wrinkle.

Filensin and phakinin form a novel type of beaded intermediate filaments and coassemble de novo in cultured cells

Abstract: The fiber cells of the eye lens possess a unique cytoskeletal system known as the "beaded-chain filaments" (BFs). BFs consist of filensin and phakinin, two recently characterized intermediate filament (IF) proteins. To examine the organization and the assembly of these heteropolymeric IFs, we have performed a series of in vitro polymerization studies and transfection experiments. Filaments assembled from purified filensin and phakinin exhibit the characteristic 19-21-nm periodicity seen in many types of IFs upon low angle rotary shadowing. However, quantitative mass-per-length (MPL) measurements indicate that filensin/phakinin filaments comprise two distinct and dissociable components: a core filament and a peripheral filament moiety. Consistent with a nonuniform organization, visualization of unfixed and unstained specimens by scanning transmission electron microscopy (STEM) reveals the the existence of a central filament which is decorated by regularly spaced 12-15-nm-diam beads. Our data suggest that the filamentous core is composed of phakinin, which exhibits a tendency to self-assemble into filament bundles, whereas the beads contain filensin/phakinin hetero-oligomers. Filensin and phakinin copolymerize and form filamentous structures when expressed transiently in cultured cells. Experiments in IF-free SW13 cells reveal that coassembly of the lens-specific proteins in vivo does not require a preexisting IF system. In epithelial MCF-7 cells de novo forming filaments appear to grow from distinct foci and organize as thick, fibrous laminae which line the plasma membrane and the nuclear envelope. However, filament assembly in CHO and SV40-transformed lens-epithelial cells (both of which are fibroblast-like) yields radial networks which codistribute with the endogenous vimentin IFs. These observations document that the filaments formed by lens-specific IF proteins are structurally distinct from ordinary cytoplasmic IFs. Furthermore, the results suggest that the spatial arrangement of filensin/phakinin filaments in vivo is subject to regulation by host-specific factors. These factors may involve cytoskeletal networks (e.g., vimentin IFs) and/or specific sites associated with the cellular membranes.

A piezoresistive carbon filament polymer-matrix composite strain sensor

Abstract: The relationship between strain and the fractional increase in electrical resistance of piezoresistive polyether-sulfone-matrix composite strain sensors was found to be much more linear and less noisy when the electrically conducting filler was 0.1 m diameter carbon filaments rather than the conventionally used 10 m diameter carbon fibers. For the fiber composite, the non-linearity manifested itself as increasing reversibly with increasing compressive strain - an effect opposite to and occurring on top of piezoresistivity. This effect was absent from the filament composite. Furthermore, the percolation threshold was lower for the filament composite than for the fiber composite. For both filament and fiber composites, became more negative as cycling progressed up to 10 cycles and then stabilized though the effect was more significant for the latter.

Fluid shear stress induces the phosphorylation of small heat shock proteins in vascular endothelial cells

Abstract: The small molecular mass heat shock protein of 27 kDa (HSP27) has been shown to influence actin filament dynamics and endothelial cell behavior in ways similar to those observed during laminar flow...

Movement actuator/sensor systems

Abstract: A movement actuator includes an elongate filament made of a flexible material, and a strip of shape memory alloy disposed on the surface of one side of the filament. The shape memory alloy is responsive to actuation signals, heat or electrical signals, for changing its shape and when its shape changes, it causes the filament to move, i.e., bend, to accommodate the change in shape of the alloy. Also included is a signal supply device for selectively applying heat signals or electrical current to the strip of shape memory alloy to cause the alloy to change its shape and cause the filament to bend. Other patterns for the shape memory alloy could be disposed on the filament to cause other kinds of movements. For example, a helical pattern of the shape memory alloy about the filament would cause the filament to twist when the helical pattern were caused to shorten or lengthen.

Ultrastructure of invertebrate muscle cell types

Abstract: The muscular cells of invertebrates can be divided into three major classes on the basis of their striation pattern: transversely striated, obliquely striated, or smooth muscle. Transversely striated muscles have either continuous or discontinuous Z lines and, thus, can be subdivided into two types respectively. Of all invertebrate muscles, the transversely striated muscle with continuous Z lines is the most similar to the vertebrate skeletal muscle and is present in arthropods, whose musculature (including the visceral muscles) only consists of this cell type. These muscles are multinucleate cells that contain myofibrils showing well-defined sarcomeres. Transversely striated muscles with discontinuous Z lines, consisting of multiple small electrondense patches, are found in the translucent portions of adductor muscles of some bivalves and in the heart muscle of the gastropods. This muscle is formed by mononucleated cells with centrally-located nuclei and a single myofibril. The obliquely striated muscle appears in nematodes, annelids, molluscs, brachiopods and chaetognathes and consists of mononucleated cells with both thick and thin myofilaments which form sarcomeres delimited by Z lines. Myofilaments are not perpendicular but oblique to the Z lines, so that both A and I bands may be seen together in each of the three spatial planes of view. Smooth muscle has been reported in coelenterates, annelids, molluscs, brachiopods and echinoderms, but is lacking in arthropods. These muscle cells have a centrally-located nucleus and abundant thin and thick myofilaments without apparent sarcomeres. The most relevant characteristics of invertebrate muscle cells are the following. The thick (myosin) myofilaments show a variable length (from 2.2 microns up to 6 microns) and width (from 14 nm up to 231 nm) and contain a central core of paramyosin, which is absent in vertebrate muscles. Thick filaments are homogenous in transversely striated muscles and either homogeneous or fusiform in the obliquely striated and smooth muscles. Thin filaments measure 6 nm in diameter. They contain tropomyosin and, only in striated muscles, also troponin. The thin/thick filament ratio varies from 3/1 to 6/1, even in smooth muscles. The plaques for filament anchorage (Z lines in striated muscles or electrondense bodies in smooth muscles) contain alpha-actinin. The striated (transversely or obliquely) muscles show long sarcomeres (up to 9 microns) and the number of thin filaments around each thick filament varies from 3 to 12, so that each thin filament is shared by two thick filaments. Z lines in the striated muscles show a variety of structures that differ from one species to another (filament bundles in nematodes, bars in annelids, small patches in molluscs, etc). Many striated muscles contain titin (connectin) and intermediate filaments and display a sarcotubular system consisting of T tubules and sarcoplasmic reticulum tubules. Both structures form dyads and, more rarely, triads. The location of T tubules as well as the configuration and distribution of sarcoplasmic reticulum vary among muscles and species. Invertebrate smooth muscle differs from that of vertebrates principally in the higher proportion and larger diameter of thick myofilaments. These may be fusiform and their size and number may vary widely among cells. These muscle cells may be classified by the characteristics of both the thick filaments and the electrondense bodies for filament anchorage.

Direct interaction of flagellin termini essential for polymorphic ability of flagellar filament

Abstract: We report the structures of flagellar filaments reconstituted from various flagellins with small terminal truncations. Flagellins from Salmonella typhimurium strains SJW1103 (wild type), SJW1660, and SJW1655 were used, which form a left-handed supercoil, the L- and R-type straight forms, respectively. Structure analyses were done by electron cryomicroscopy and helical image reconstruction with a help of x-ray fiber diffraction for determining precise helical symmetries. Truncation of either terminal region, irrespective of the original flagellin species, results in a straight filament having a helical symmetry distinct either from the L- or R-type. This filament structure is named Lt-type. Although the local subunit packing is similar in all three types, a close comparison shows that the Lt-type packing is almost identical to the R-type but distinct from the L-type, which demonstrates the strong two-state preference of the subunit interactions. The structure clearly suggests that both termini are located in the inner tube of the concentric double-tubular structure of the filament core, and their proper interaction is responsible for the correct folding of fairly large terminal regions that form the inner tube. The double tubular structure appears to be essential for the polymorphic ability of flagellar filaments, which is required for the swimming-tumbling of bacterial taxis.