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J Bordas

Publications and source records attributed to J Bordas.

At least 19 recordsLinked to original sources
PubMed2001

Axial disposition of myosin heads in isometrically contracting muscles.

Meridional x-ray diffraction diagrams, recorded with high angular resolution, from muscles contracting at the plateau of isometric tension show that the myosin diffraction orders are clusters of peaks. These clusters are due to pronounced interference effects between the myosin diffracting units on either side of the M-line. A theoretical analysis based on the polarity of the myosin (and actin) filaments shows that it is possible to extract phase information from which the axial disposition of the myosin heads can be determined. The results show that each head in a crown pair has a distinct structural disposition. It appears that only one of the heads in the pair stereospecifically interacts with the thin filament at any one time.

Actins↗
PubMed1999

Extensibility and symmetry of actin filaments in contracting muscles.

When isometrically contracting muscles are subjected to a quick release followed by a shortening ramp of appropriate speed (V(o)), tension decays from its value at the isometric plateau (P(o)) to <0. 05 P(o) with the same time course as the quick part of the release; thereafter, tension remains at a negligible level for the duration of the shortening ramp. X-ray diffraction data obtained under these conditions provide evidence that 1) at V(o) very few heads form an actomyosin complex, while the number of heads doing so at P(o) is significant; 2) relative to rest the actin filament at V(o) is approximately 0.12% shorter and more twisted, while it is approximately 0.3% longer and less twisted at P(o); and 3) the myosin heads attaching to actin during force development do so against a thin filament compliance of at least 0.646 +/- 0.046% nm per P(o).

Actins↗
PubMed1998

The susceptibility of pure tubulin to high magnetic fields: a magnetic birefringence and x-ray fiber diffraction study.

The orientational behavior of microtubules assembled in strong magnetic fields has been studied. It is shown that when microtubules are assembled in a magnetic field, they align with their long axis parallel to the magnetic field. The effect of several parameters known to affect the microtubule assembly are investigated with respect to their effect on the final degree of alignment. Aligned samples of hydrated microtubules suitable for low-resolution x-ray fiber diffraction experiments have been produced, and the results obtained from the fiber diffraction experiments have been compared with the magnetic birefringence experiments. Comparisons with earlier fiber diffraction work and small-angle x-ray solution scattering experiments have been made.

Animals↗
PubMed1998

Randomized trial of adjuvant chemotherapy with mitomycin plus ftorafur versus mitomycin alone in resected locally advanced gastric cancer.

PURPOSE: We performed a clinical trial to determine whether postoperative adjuvant chemotherapy with two drugs versus one drug could prolong survival. PATIENTS AND METHODS: From 1985 to 1996, 85 patients with completely resected locally advanced gastric cancer were enrolled. The subjects were randomized into two treatment groups, as follows: mitomycin (MMC) 10 to 20 mg/m2 intravenously (i.v.) on day 1 every 6 weeks plus ftorafur (FT) 500 mg/m2/d for 36 consecutive days; or MMC alone, 10 to 20 mg/m2 i.v. every 6 weeks. All courses were repeated four times. RESULTS: After a median follow-up duration of 62 months, the overall 5-year survival rates were 67% for the MMC-FT group versus 44% for the MMC group (P = .04). Subgroup analysis to compare survival curves using the method of Mantel-Cox showed survival rates significantly in favor of the MMC-FT group in the subsets of patients with node-negative disease (P = .01) and those whose disease was stage IB or II (P = .008). CONCLUSION: Significantly better survival results were observed for MMC-FT versus MMC alone. Subset analysis suggest a strong benefit in patients with node-negative and early-stage resected gastric cancer.

Adenocarcinoma↗
PubMed1996

Small segmental rearrangements in the myosin head can explain force generation in muscle.

Poisson-Boltzmann calculations of the distribution of electrostatic potentials around an actin filament in physiological-strength solutions show that negative isopotential surfaces protrude into the solvent. Each protrusion follows the actin two-start helix and is located on the sites implicated in the formation of the actomyosin complex. Molecular dynamic calculations on the S1 portion of the myosin molecule indicate that in the presence of ATP the crystallographically invisible loops (comprising residues 624-649 and 564-579) remain on the surface, whereas in the absence of ATP they can move toward the actin-binding sites and experience electrostatic forces that range from 1 to 10 pN. The molecular dynamics calculations also suggest that during the ATP cycle there exist at least three states of electrostatic interactions between the loops and actin. Every time a new interaction is formed, the strain in the myosin head increases and the energy of the complex decreases by 2kT to 5kT. This can explain muscular contraction in terms of a Huxley-Simmons-type mechanism, while requiring only rearrangements of small mobile S1 segments rather than the large shape changes in the myosin molecule postulated by the conventional tilting head model.

Actins↗
PubMed1996

Structural intermediates in the assembly of taxoid-induced microtubules and GDP-tubulin double rings: time-resolved X-ray scattering.

We have studied the self-association reactions of purified GDP-liganded tubulin into double rings and taxoid-induced microtubules, employing synchrotron time-resolved x-ray solution scattering. The experimental scattering profiles have been interpreted by reference to the known scattering profiles to 3 nm resolution and to the low-resolution structures of the tubulin dimer, tubulin double rings, and microtubules, and by comparison with oligomer models and model mixtures. The time courses of the scattering bands corresponding to the different structural features were monitored during the assembly reactions under varying biochemical conditions. GDP-tubulin essentially stays as a dimer at low Mg(2+) ion activity, in either the absence or presence of taxoid. Upon addition of the divalent cations, it associates into either double-ring aggregates or taxoid-induced microtubules by different pathways. Both processes have the formation of small linear (short protofilament-like) tubulin oligomers in common. Tubulin double-ring aggregate formation, which is shown by x-ray scattering to be favored in the GDP- versus the GTP-liganded protein, can actually block microtubule assembly. The tubulin self-association leading to double rings, as determined by sedimentation velocity, is endothermic. The formation of the double-ring aggregates from oligomers, which involves additional intermolecular contacts, is exothermic, as shown by x-ray and light scattering. Microtubule assembly can be initiated from GDP-tubulin dimers or oligomers. Under fast polymerization conditions, after a short lag time, open taxoid-induced microtubular sheets have been clearly detected (monitored by the central scattering and the maximum corresponding to the J(n) Bessel function), which slowly close into microtubules (monitored by the appearance of their characteristic J(0), J(3), and J (n) - (3) Bessel function maxima). This provides direct evidence for the bidimensional assembly of taxoid-induced microtubule polymers in solution and argues against helical growth. The rate of microtubule formation was increased by the same factors known to enhance taxoid-induced microtubule stability. The results suggest that taxoids induce the accretion of the existing Mg(2+)-induced GDP-tubulin oligomers, thus forming small bidimensional polymers that are necessary to nucleate the microtubular sheets, possibly by binding to or modifying the lateral interaction sites between tubulin dimers.

Animals↗
PubMed1995

The effect of temperature on the structure of vinblastine-induced polymers of purified tubulin: detection of a reversible conformational change.

Addition of the antimitotic drug vinblastine to solutions of purified tubulin induces the formation of helical polymers whose structure and type of aggregation is determined by the concentration of magnesium. While paracrystalline arrangements of single coils are observed at low concentrations of the ion, for concentrations higher than 6 mM free double-coiled spirals are obtained, which are indistinguishable from those obtained in the presence of microtubule-associated proteins (MAPs). This result is consistent with a similar effect of magnesium and MAPs in neutralizing negative charges on the tubulin molecule and so allowing for lateral contacts between protofilaments. The effects that temperature has on the structure of both types of polymers, free spirals or paracrystals, have been monitored using time-resolved X-ray solution scattering. This study shows that a temperature increase: (1) affects the length and lateral aggregation of the spirals in the paracrystalline sample; (2) induces a reversible increase of the helical pitch in both types of polymers that closely follows the temperature change; (3) produces an irreversible aggregation of some of the protein in both types of polymers; and (4) can induce a reversible transformation from one type of structure to the other when the concentration of Mg2+ is in the boundary between the two ranges. We suggest that the changes in pitch are due to a temperature-induced conformational change of the tubulin molecule. This effect may be related to the structural modifications that result in the temperature-induced assembly of microtubules in vitro under normal conditions of assembly.

Animals↗
PubMed1995

X-ray evidence that in contracting live frog muscles there exist two distinct populations of myosin heads.

Using synchrotron radiation and whole muscles, 2 ms time-resolved x-ray diffraction patterns were recorded at 8 degrees C. The results show that in both isotonic and isometric contractions, as well as in length changes imposed at maximum tension [Po], the meridional third myosin layer line consists of two distinct reflections with different intensities and spacings that measure approximately 14,623 and 14,412 nm at Po. Although the intensity behavior of the two reflections is strikingly different during quick releases, it is very similar during stretches. Study of the time courses indicates that myosin heads diffracting at Po with the approximately 14.623 nm periodicity are actively involved in tension production. Those diffracting at Po with the periodicity of approximately 14.412 nm appear not be associated with tension production during isometric contraction and releases, but the results suggest that they are recruited during stretches and here contribute to tension production. Our most important conclusion is that under all conditions of contraction we have investigated there exist two populations of myosin heads, each with a well defined axial disposition and configuration.

Animals↗
PubMed1994

Solution structure of Taxotere-induced microtubules to 3-nm resolution. The change in protofilament number is linked to the binding of the taxol side chain.

The synchrotron x-ray solution scattering profiles of microtubules assembled from purified GDP- or GTP-tubulin with the antitumor drug docetaxel (Taxotere) are consistent with identical non-globular alpha and beta-tubulin monomers ordered within the known surface lattice of microtubules, with a center to center lateral spacing of 5.7 +/- 0.1 nm. The higher angle part of the scattering profile, and therefore the substructure of the microtubule wall is identical in Taxotere- and Taxol-induced microtubules, to the resolution of the measurements. However, Taxotere-induced microtubules have a mean diameter of 24.2 +/- 0.4 nm, which is 1.12 +/- 0.01 times larger than that of paclitaxel (Taxol) induced microtubules. The population of Taxotere microtubules has on average 13.4 protofilaments, which is similar to control microtubules assembled with glycerol but is in marked contrast with Taxol-induced microtubules, which have on average 12 protofilaments under identical solution conditions. Model populations of Taxotere and Taxol microtubules with the distributions of protofilament numbers determined by electron microscopy reproduce the positions and approximate intensities of the experimental x-ray scattering data. Comparison of the structures and activities of both taxoids strongly suggests that the change of the more frequent lateral bond angle between tubulin molecules from 152.3 degrees (13-protofilament microtubules) to 150 degrees (12-protofilament microtubules) is linked to the binding of the side chain of Taxol. Optimal microtubule formation is obtained with unitary Taxotere to tubulin heterodimer ratio; however, ligand molecules in excess over tubulin dimers cause a loss of cylindrical scattering features, consistent with microtubule opening. The results are compatible with the observed biochemical and thermodynamic properties of this ligand-induced microtubule assembly system and also with the simple working hypothesis that taxoids would bind between adjacent microtubule protofilaments.

Docetaxel↗
PubMed1994

X-ray diffraction studies of whole rat heart during anoxic perfusion.

Equatorial x-ray reflections were recorded from the ventricular region of whole rat heart, during the transition from normoxia to anoxia. The intensity ratio of the 1.0 and 1.1 equatorial reflections (I 1.0/I 1.1) was 2.96 during normoxic perfusion, decreasing to 0.37 after 115 minutes of anoxic perfusion and closely parallel those reported between partially dissected relaxed and rigor vertebrate heart muscle (1.2). The peak positions of these reflections both increased by ca. 5% during anoxic perfusion indicative of a lateral expansion in the sarcomere filament lattice. These results indicate that the process of anoxia leads to the condition of rigor in which the majority of the myosin cross-bridges bind to the thin filament. Using this technique global changes in whole heart structure can be studied, and due to the ease of perfusion of the heart, biochemical and physiological problems may be investigated in relation to the structure of the heart as a whole. This may be of clinical interest, particularly in terms of investigations into organ preservation and transplantation. This is believed to be the first occasion where intact muscular organs have been studied in this way.

Animals↗
PubMed1994

Solution structure of GDP-tubulin double rings to 3 nm resolution and comparison with microtubules.

GDP liganded tubulin, which is inactive in microtubule assembly, polymerizes into rings more readily than the active GTP liganded protein. The structure of double rings made of highly purified GDP-tubulin has been characterized to 3 nm resolution with synchrotron X-ray solution scattering. The scattering profile has characteristic maxima due to closely packed double rings of 38 nm mean diameter, with a 5.5 nm mean spacing between the rings, and a 4.2 nm centre-to-centre spacing between non-globular tubulin monomers within both rings. There are probably 24 and 32 monomers in the inner and outer ring, respectively, and the double ring population is more than 75% homogeneous in size. Comparison of this double ring structure to the lattice of tubulin molecules in microtubules indicates that the tubulin rings are equivalent to pairs of protofilament segments curved tangentially to the microtubule surface, with bending angles of 30 degrees and 22.5 degrees per tubulin alpha beta dimer. When the rings are modelled employing the same non-globular tubulin monomer as in microtubules, the best computer fitted scattering profiles correspond to monomer orientations equivalent to two microtubule protofilaments coiled sideways, with same or opposite polarity. Rings constitute the equilibrium assembly state of GDP-tubulin, which is tensioned inside microtubules after GTP hydrolysis, causing their functional instability. In analogy with other nucleotide binding proteins, the inactive/active structural switch of tubulin is induced by the binding of the gamma phosphate and a coordinated Mg ion. It should involve domain rearrangements which cancel the bending of the tubulin dimer in the ring structure.

Guanosine Diphosphate↗
PubMed1994

Time-resolved X-ray diffraction studies of myosin head movements in live frog sartorius muscle during isometric and isotonic contractions.

Using the facilities at the Daresbury Synchrotron Radiation Source, meridional diffraction patterns of muscles at ca 8 degrees C were recorded with a time resolution of 2 or 4 ms. In isometric contractions tetanic peak tension (P0) is reached in ca 400 ms. Under such conditions, following stimulation from rest, the timing of changes in the major reflections (the 38.2 nm troponin reflection, and the 21.5 and 14.34/14.58 nm myosin reflections) can be explained in terms of four types of time courses: K1, K2, K3 and K4. The onset of K1 occurs immediately after stimulation, but that of K2, K3 and K4 is delayed by a latent period of ca 16 ms. Relative to the end of their own latent periods the half-times for K1, K2, K3 and K4 are 14-16, 16, 32 and 52 ms, respectively. In half-times, K1, K2, K3 lead tension rise by 52, 36 and 20 ms, respectively. K4 parallels the time course of tension rise. From an analysis of the data we conclude that K1 reflects thin filament activation which involves the troponin system; K2 arises from an order-disorder transition during which the register between the filaments is lost; K3 is due to the formation of an acto-myosin complex which (at P0) causes 70% or more of the heads to diffract with actin-based periodicities; and K4 is caused by a change in the axial orientation of the myosin heads (relative to thin filament axis) which is estimated to be from 65-70 degrees at rest to ca 90 degrees at P0. Isotonic contraction experiments showed that during shortening under a load of ca 0.27 P0, at least 85% of the heads (relative to those forming an acto-myosin complex at P0) diffract with actin-based periodicities, whilst their axial orientation does not change from that at rest. During shortening under a negligible load, at most 5-10% of the heads (relative to those forming an acto-myosin complex at P0) diffract with actin-based periodicities, and their axial orientation also remains the same as that at rest. This suggests that in isometric contractions the change in axial orientation is not the cause of active tension production, but rather the result of it. Analysis of the data reveals that independent of load, the extent of asynchronous axial motions executed by most of the cycling heads is no more than 0.5-0.65 nm greater than at rest.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1993

Two-dimensional time-resolved X-ray diffraction studies of live isometrically contracting frog sartorius muscle.

Results were obtained from contracting frog muscles by collecting high quality time-resolved, two-dimensional, X-ray diffraction patterns at the British Synchrotron Radiation Source (SERC, Daresbury, Laboratory). The structural transitions associated with isometric tension generation were recorded under conditions in which the three-dimensional order characteristic of the rest state is either present or absent. In both cases, new layer lines appear during tension generation, subsequent to changes from activation events in the thin filaments. Compared with the 'decorated' actin layer lines of the rigor state, the spacings of the new layer lines are similar whereas their intensities differ substantially. We conclude that in contracting muscle an actomyosin complex is formed whose structure is not like that in rigor, although it is possible that the interacting sites are the same. Transition from rest to plateau of tension is accompanied by approximately 1.6% increase in the axial spacing of the myosin layer lines. This is explained as arising from axial disposition of the interacting myosin heads in the actomyosin complex. Model calculations are presented which support this view. We argue that in a situation where an actomyosin complex is formed during contraction, one cannot describe the diffraction features as being either thick or thin filament based. Accordingly, the layer lines seen during tension generation are referred to as actomyosin layer lines. It is shown that these layer lines can be indexed as submultiples of a minimum axial repeat of approximately 218.7 nm. After lattice disorder effects are taken into account, the intensity increases on the 15th and 21st AM layer lines at spacings of approximately 14.58 and 10.4 nm respectively, show the same time course as tension rise. However, the time course of the intensity increase of the other actomyosin layer lines and of the spacing change (which is the same for both phenomena) shows a substantial lead over tension rise. These findings suggest that the actomyosin complex formed prior to tension rise is a non-tension-generating state and that this is followed by a transition of the complex to a tension-generating state. The intensity increase in the 15th actomyosin layer line, which parallels tension rise, can be accounted for assuming that in the tension-generating state the attached heads adopt (axially) a more perpendicular orientation with respect to the muscle axis than is seen at rest or in the non-tension-generating state. This suggests the existence of at least two structurally distinct interacting myosin head conformations.(ABSTRACT TRUNCATED AT 400 WORDS)

Actomyosin↗
PubMed1992

Low resolution structure of microtubules in solution. Synchrotron X-ray scattering and electron microscopy of taxol-induced microtubules assembled from purified tubulin in comparison with glycerol and MAP-induced microtubules.

The structure of microtubules has been characterized to 3 nm resolution employing time-resolved X-ray scattering. This has revealed detailed structural features of microtubules not observed before in solution. The polymerization of highly purified tubulin, induced by the antitumour drug taxol, has been employed as a microtubule model system. This assembly reaction requires Mg2+, is optimal at a 1:1 taxol to tubulin heterodimer molar ratio, proceeds with GTP or GDP and is intrinsically reversible. The X-ray scattering profiles are consistent with identical non-globular alpha and beta-tubulin monomers ordered within the known helical surface lattice of microtubules. Purified tubulin-taxol microtubules have a smaller mean diameter (approx. 22 nm) than those induced by microtubule associated proteins or glycerol (approx. 24 nm), but nearly identical wall substructure to the resolution of the measurements. This is because the majority of the former consist of only 12 protofilaments instead of the typical 13 protofilaments, as confirmed by electron microscopy of thin-sectioned, negatively stained and ice-embedded taxol microtubules. It may be concluded that taxol induces a slight reduction of the lateral contact curvature between tubulin monomers. The main fringe pattern observed in cryo-electron micrographs is consistent with a simple 12 protofilament 3-start skewed lattice model. Cylindrical closure of this lattice can be achieved by tilting the lattice 0.8 degrees with respect to the microtubule axis. The closure implies a discontinuity in the type of lateral contacts between the tubulin monomers (regardless of whether these are of the -alpha-beta- or the -alpha-alpha-/-beta-beta- type), which indicates that lateral contacts and the subunit specificity of taxol binding are, to a large degree, equivalent.

Alkaloids↗
PubMed1992

Chronic systemic candidiasis in acute leukemia.

In the past few years a new syndrome of invasive Candida infection, the so-called hepatosplenic or chronic systemic candidiasis (CSC), has been recognized with increasing frequency in neutropenic patients. From January 1985 to December 1990, ten of 305 acute leukemia (AL) patients treated at our institution were diagnosed as having CSC. In contrast, during the same period this type of Candida infection was not observed in any patient with hematological diseases other than AL treated in our center, including 277 patients who underwent bone marrow transplantation. All patients with CSC had fever and hepatomegaly, and five complained of abdominal pain. Seven patients had neutrophilic leukocytosis and six an increased serum alkaline phosphatase activity. Abdominal computed tomography and ultrasound study showed typical lesions in eight and seven patients, respectively. In four patients a laparoscopy-guided needle liver biopsy displayed yellowish nodules on the liver surface, and the histologic study revealed large granulomas with yeasts and pseudohyphae. All patients were given amphotericin B (mean: 4.6 g, range: 1-12.5 g) and 5-fluorocytosine, and five received fluconazole. No patient died as a direct consequence of CSC and in six the infection resolved. Finally, once controlled, the infectious complication did not preclude subsequent intensive antileukemic therapy, including bone marrow transplantation.

Adolescent↗
PubMed1992

Structure of the myosin head in solution and the effect of light chain 2 removal.

Structural properties of rabbit skeletal myosin head (S1) and the influence of the DTNB light chain (LC2) on the size and shape of myosin heads in solution were investigated by small angle x-ray scattering. The LC2 deficient myosin head, S1 (-LC2), and the S1 containing LC2 light chain, S1 (+LC2) were studied in parallel. The respective values of the radius of gyration were found to be (40.2 +/- 0.5) A and (46.7 +/- 1) A, while the maximum dimension was (190 +/- 15) A for both species. The large difference between the two Rg values suggest that LC2 is located close to one extremity of the myosin head, in agreement with most electron microscopy observations. All models derived from the x-ray scattering pattern of the native myosin head share a common overall morphology, showing two main regions, an asymmetric globular portion which tapers smoothly into a thinner domain of roughly equivalent length making an angle of approximately 60 degrees, with a contour length of approximately 210 A.

Animals↗