PubMed Health⌕ Search

Biomedical subjects

E Grazi

Publications and source records attributed to E Grazi.

At least 55 records · Page 3Linked to original sources
PubMed1988

On the interaction between xanthine oxidase and actin.

Xanthine oxidase increases the rate of actin polymerization. This occurs at oxidase concentrations as low as 40 nM provided the concentration of the polymerizing agent is low (0.5 mM MgCl2). In the presence of 0.1 M KCl plus 1 mM MgCl2 as the polymerizing agents, xanthine oxidase does not affect the rate of the polymerization but increases significantly the rate of the conversion of F(ATP)actin into F(ADP.Pi)actin and probably also the rate of the orthophosphate release.

Actins↗
PubMed1987

Kinetic heterogeneity of F-actin polymers. Further evidence that the elongation reaction may occur through condensation of the actin filaments with small aggregates.

We have shown that F-actin, polymerized in 50 mM-KCl at 20 degrees C and pH 8.0, can be resolved by centrifugation into two polymer populations, which differ morphologically as well as kinetically. The first population represents about 10% of the overall polymer and is composed of small amorphous aggregates. It rapidly exchanges the bound nucleotide with free ATP in the medium, either directly or through the monomers. The second population is composed of long actin filaments. These are labelled by free ATP in the medium only through condensation with labelled small amorphous aggregates.

Actins↗
PubMed1987

On an Mg2+-dependent interaction of actin with glyceraldehyde-phosphate dehydrogenase. The fundamental role of KCl in the organization of F-actin.

In the presence of Mg2+, the formation of actin filaments is hindered by glyceraldehyde-3-phosphate dehydrogenase. This effect, which increases with the square of Mg2+ concentration, is counteracted by 0.15 M KCl. Thus KCl, at concentrations found in the intracellular compartment, appears to be strictly required for the correct formation of actin filaments in all tissues in which the glyceraldehyde-phosphate dehydrogenase concentration is high.

Actins↗
PubMed1987

The experimental basis of some recent hypotheses on the mechanism of the polymerization of actin: a reappraisal.

We have studied the effect of sonication on the fluorescence of N-(1-pyrenyl)iodoacetamide-labeled F-actin as well as of native actin-pyrenyl-actin mixed oligomers in which the subunits were covalently attached to each other by phenylenebismaleimide. In both cases the fluorescence of the solution was largely decreased by sonication. We have found that this effect is due (a) to a 20-30% decrease of the specific fluorescence of the polymers. These results question the validity of the novel mechanism for the polymerization of actin recently proposed (D. Pantaloni et al. (1984) J. Biol. Chem. 259, 6274-6283). In these studies, in fact, the implicit assumption was made that the quenching of the fluorescence of the solution under sonication was due exclusively to the conversion of F-actin into G-actin.

Actins↗
PubMed1987

Fructose bisphosphate aldolase from rabbit muscle. A jump in the van't Hoff plot accompanies the onset of half of the sites' reactivity.

In 40% ethylene glycol, gamma/2 = 0.11 and pH* 8.2, fructose 1,6-bisphosphate aldolase from rabbit muscle undergoes a transition: above 3 degrees C it displays 4 equivalent dihydroxyacetone phosphate binding sites, below -1 degree C the sites decrease to 2. The dissociation constant of the aldolase-dihydroxyacetone phosphate complex decreases from 10 microM at 3 degrees C to 2.65 microM at -1 degree C, its van't Hoff plot being linear between -1 degree C and -13 degrees C. The rate of the detritiation of the aldolase-(3S)-[3-3H]dihydroxyacetone phosphate complex is strongly influenced by temperature. In 40% ethylene glycol, gamma/2 = 0.01 and pH* 8.2, the apparent rate constant is 7.6 sec-1 at -5 degrees C and 0.012 sec-1 at -24 degrees C. The Arrhenius plot is linear between -5 degrees C and -24 degrees C.

Animals↗
PubMed1986

An ADPase from sarcoplasmic reticulum of rabbit muscle cleaves ADP bound to F-actin.

We have found that sarcoplasmic reticulum from rabbit muscle contains an ADPase which cleaves ADP bound to F-actin. The interaction is not of the simple Michaelis-Menten type, the order of the reaction being larger than the first. A possible explanation of this behaviour could be that ADPase binds to two adjacent actin monomers with a preferred orientation thus cleaving preferentially the nucleotide of one of the two monomers.

Actins↗
PubMed1985

Effects of temperature on actin polymerized by Ca2+. Direct evidence of fragmentation.

When the temperature is lowered from 20 to 4 degrees C, the specific viscosity of actin polymerized in the presence of either 4 mM-CaCl2 or 2 mM-MgCl2, but not of actin polymerized in the presence of 90 mM-KCl, is decreased by 50% in the absence of free ATP. Addition of ATP restores the viscosity of the actin polymerized by Mg2+, but not that of actin polymerized by Ca2+, to the original value. The effect of temperature on actin polymerized in the presence of Ca2+ is due to (a) polymer-into-monomer conversion, (b) latero-lateral aggregation of filaments, and (c) fragmentation of the filaments. Fragmentation, as demonstrated by fractional centrifugation and electron microscopy, was the most important of these.

Actins↗
PubMed1984

54Mn2+ as a tracer of the polymerization of actin. Intermediate oligomers condense to give F-actin.

Mg2+, at submicromolar concentrations, is needed for the nucleation of actin [Maruyama (1981) J. Biol. Chem. 256, 1060-1062]. I show here that Mn2+ fulfils the same function. It binds to oligomers present in the ATP-G-actin solutions with a ratio of 2-3 Mn2+ ions per 100 actin monomers and with an association constant of 0.66 X 10(10) M-1 at pH 8.2 at 25 degrees C. The time course of the binding of Mn2+ to polymerizing actin is not affected by the initial concentration of the protein. Analysis of the distribution of the binding shows that, both in the large oligomeric species and in the polymers, 1 Mn2+ ion is bound for every 14-25 actin monomers, whereas in the smaller oligomeric species 1 Mn2+ ion is bound for every 4 actin monomers. The proposal is made that Mn2+ stabilizes actin nuclei and decreases the concentration of the monomers at the steady state. It is also proposed that, at least in some experimental conditions, the direct condensation of oligomers of intermediate length is an effective mechanism of F-actin formation.

Actins↗
PubMed1984

Characterization of the ATP-G-actin aggregates formed at low potassium chloride concentration.

The ATP-G-actin aggregates formed by incubation of ATP-G-actin in 7.5 mM-KCl were characterized by electron-microscopical observation, by high-pressure liquid chromatography and by the study of the 1,N6-etheno-ATP-ATP exchange reaction between the free and the actin-bound nucleotide. In 30 mM-KCl the initial rate of the reduced-viscosity increase is found to be directly related to the amount of the aggregates formed in the course of the preincubation in 7.5 mM-KCl.

Actins↗
PubMed1984

Fructose-1,6-bisphosphate aldolase from rabbit muscle: different catalytic behavior of the dihydroxyacetone phosphate binding sites at low temperature.

The equivalence of the four dihydroxyacetone phosphate binding sites of aldolase was abolished by lowering the temperature. At pH 6.2 and -13 degrees C, four binding sites were detected by gel filtration; two sites with a Kdiss less than or equal to 0.1 microM, and a second set of sites with a Kdiss = 4 microM. The alteration of the binding was accompanied by the alteration of the catalytic activity. The low-affinity sites were incapable of catalyzing the cleavage of the (3S) C-H bond of dihydroxyacetone phosphate, and form only the ketimine phosphate intermediate. The high-affinity sites were still able to cleave the (3S) C-H bond of dihydroxyacetone phosphate; however, the eneamine phosphate intermediate formed was almost fully converted into the eneamine-aldehyde . . . phosphate intermediate, which was the prevailing species at the equilibrium. The mechanism of the half-of-the sites reactivity of aldolase at low temperature has been explained and the nonequivalence of sites in promoting catalysis has been utilized to dissect and characterize the individual partial reactions of the enzyme. In the course of these studies it has been shown that the rate of hydration-dehydration of dihydroxyacetone phosphate at -24 degrees C was too slow to measure.

Animals↗
PubMed1984

A mechanism for the selective preservation of homogeneous. F(ATP) actin.

The cold-induced depolymerization of F(ADP) actin, coupled with the repolymerization by ATP, was employed to study the formation of F(ATP) actin. It is proposed that the cold-induced lability of F(ADP) actin, together with the spontaneous fragmentation of the filament, provides a mechanism for the selective preservation of homogeneous F(ATP) actin.

Actins↗
PubMed1983

Fructose-1,6-bisphosphate aldolase from rabbit muscle. Kinetic resolution of the enamine phosphate from the enamine-aldehyde intermediate at low temperature.

At or below -12 degrees C and in the presence of 40% ethylene glycol, only two out of the four dihydroxyacetone phosphate binding sites of aldolase are catalytically active. At these same temperatures and at pH* 8.3, the equilibrium between the pre-enamine and the enamine plus the post-enamine intermediates is largely shifted in favor of the latter. The enamine phosphate and the enamine-aldehyde phosphate intermediates have been resolved by studying the rate of their formation at -13 degrees C and pH* 5.28 and the trapping by DL-glyceraldehyde 3-phosphate at -24 degrees C and pH* 5.24.

Aldehydes↗
PubMed1983

The polymerization of actin. A study of the nucleation reaction.

We compared the properties of the nuclei that accumulate in 7.5 mM-KCl in ATP-G-actin solutions and of the oligomers that are formed by sonication of either G-actin or F-actin. We found that the ability of the above species to prime the polymerization of actin decays with different rates. The nuclei are stable in 7.5 mM-KCl (they decay with a rate constant of 1.5 X 10(-3) s -1 at pH 7.8 at 22 degrees C in the absence of KCl). The oligomers formed by sonication of either G-actin or F-actin, once the sonication is stopped, revert to simpler structures or evolve into F-actin, depending on the KCl concentration in which they are kept. In 10.5 mM-KCl at pH 7.8 at 22 degrees C their priming ability decays with a rate constant of 6 X 10(-3) s -1. We propose that the nuclei that form spontaneously in 7.5 mM-KCl are not directly susceptible to elongation. They must first be converted into activated nuclei, which exist in very low concentration at the steady state. The activated nuclei are directly susceptible to elongation, they have a short life and they decay rapidly into the ground state unless the elongation reaction occurs. Sonication displaces the steady-state concentration in favour of the activated state.

Actins↗
PubMed1982

Mechano-chemical energy transduction in biological systems. The effect of mechanical stimulation on the polymerization of actin: a kinetic study.

Mechanical stimulation (forced circulation in narrow tubing) accelerates as much as 10-fold the rate of polymerization of actin. The increase in the rate is proportional to the intensity of the stimulation for flow rates between 0 and 3 cm/s. This supports the hypothesis that a statistical factor (the orientation of the flowing particles) is influenced by the flow. Comparison of the kinetics of the polymerization of resting and of mechanically stimulated actin solutions shows that both the nucleation and the elongation steps are accelerated. It is thus concluded that flow orients not only the oligomeric structures but also the actin monomers. The elongation reaction, also in the flow-stimulated samples, occurs always by the addition of ATP--G-actin (or ATP-containing oligomers) and not by the fusion of ADP-containing oligomeric structures.

Actins↗