Characterization of the binding of paylean and DNA by fluorescence,UV spectroscopy and molecular docking techniques

The interaction of paylean (PL) with calf thymus DNA (ctDNA) was investigated using fluorescence spectroscopy, UV absorption, melting studies, ionic strength, viscosity experiments and molecular docking under simulated physiological conditions. Values for the binding constant K_a between PL and DNA were 5.11 × 10³, 2.74 × 10³ and 1.74 × 10³ L mol^–1 at 19, 29 and 39°C respectively. DNA quenched the intrinsic fluorescence of PL via a static quenching procedure as shown from Stern–Volmer plots. The relative viscosity and the melting temperature of DNA were basically unchanged in the presence of PL. The fluorescence intensity of PL–DNA decreased with increasing ionic strength. The value of K_a for PL with double‐stranded DNA (dsDNA) was larger than that for PL with single‐stranded DNA (ssDNA). All the results revealed that the binding mode was groove binding, and molecular docking further indicated that PL was preferentially bonded to A–T‐rich regions of DNA. The values for ΔH, ΔS and ΔG suggested that van der Waals forces or hydrogen bonding might be the main acting forces between PL and DNA. The binding distance was determined to be 3.37 nm based on the theory of Förster energy transference, which indicated that a non‐radiation energy transfer process occurred. Copyright © 2015 John Wiley & Sons, Ltd.     

Activation of tubulinyl-tyrosine carboxypeptidase by spermine,spermidine and putrescine

Spermine, spermidine and putrescine produce dose dependent stimulation of the invitro tubulinyl-tyrosine carboxypeptidase. Maximal stimulation was obtained with spermine, spermidine or putrescine at 0.06 mM, 1 mM and 6 mM, respectively. At higher concentrations, the enzyme activity was inhibited. The enzyme was also activated by Mg⁺⁺; the concentration formaximal effect was 4–6 mM. The stimulation produced by optimal concentration of each amine was unaffected by Mg⁺⁺ up to 2 mM; higher concentration of Mg⁺⁺ showed inhibitory effect. At optimal Mg⁺⁺ concentration, the carboxypeptidase activity was inhibited by increasing amine concentration. The amines at 0.5 or 5 mM did not produce any effect on the incorporation of tyrosine catalyzed by tubulin tyrosine ligase.     

Influence of exogenous ions on the events of maturation in Rana pipiens oocytes

Full-grown ovarian oocytes removed from non-hormone-treated Rana pipiens females exhibit a low level of protein synthesis, the rate of which is dependent upon the ionic environment. The highest rates of protein synthesis in these oocytes are obtained in media containing either a divalent cation (Ca⁺⁺ or Mg⁺⁺) or high levels of K⁺. The dependence of protein synthesis on ionic environment persists through about the first 18-24 hours of maturation (at 18°C). Normal maturation of oocytes in vitro also has specific ionic requirements for the first 24 hours. In this case, the process requires high ionic strength (T/2 = 1.0-1.2) and divalent cations. The kinetics of K⁺ exchange suggest that K⁺ exists in the ovarian oocyte in two compartments; one in equilibrium with the exogenous medium and freely exchangeable; the other in equilibrium with the exogenous medium and freely exchangeable; the other in equilibrium with the first internal compartment and only very slowly exchangeable. The slowly exchangeable (bound) compartment contains about 95% of all endogenous K⁺. In hormone stimulated oocytes, the kinetics of K⁺ exchange are essentially the same. Oocyte adaptation to ionic environment is discussed as a possible regulatory mechanism during maturation.     

Equilibrium dialysis studies of polyamine binding to DNA

Binding constants and binding site sizes for the interactions of the polyamines spermine (+4), spermidine (+3), and putrecine (+2) with helical DNA have been determined as a function of ionic conditions and temperature by equilibrium dialysis using ¹⁴C-labeled polyamines. In addition, competition equilibrium dialysis has been used to determine binding parameters for the divalent cations putrescine and Mg²⁺ from the competitive effect of these ions on the binding of spermine or spermidine. In all cases, the logarithm of the binding constant (log K_obs) varies linearly with the logarithm of the monovalent salt concentration; the slopes d log K_obs/d log[NaCl] are proportional to the valence of the ligand, and values of the extrapolated binding constants at 1M NaCl obtained from the intercepts are small (of order 1–10M^?1). In those cases examined, K_obs is insensitive to temperature; the free energy of binding is predominantly entropic. Consequently, polymines as DNA-binding ligands behave analogously to the oligolysìnes investigated previously [cf. Record, Lohman & de Haseth (1976) J. Mol. Biol. 107 , 145–158; Lohman, de Haseth & Record (1980) Biochemistry 19 , 3522–3530]. The interactions of these oligocations with DNA are predominantly electrostatic and are driven by the release of thermodynamically bound electrolyte ions from the vicinity of the DNA. The extent to which these oligocations are localized at individual phosphate binding sites or delocalized on the DNA molecule is currently not known.     

Na<Superscript>+</Superscript>,K<Superscript>+</Superscript>-ATPase Activity in the Posterior Gills of the Blue Crab, <Emphasis Type="Italic">Callinectes ornatus</Emphasis> (Decapoda,Brachyura): Modulation of ATP Hydrolysis by the Biogenic Amines Spermidine and Spermine

We investigated the effect of the exogenous polyamines spermine, spermidine and putrescine on modulation by ATP, K⁺, Na⁺, NH₄ ⁺ and Mg²⁺ and on inhibition by ouabain of posterior gill microsomal Na⁺,K⁺-ATPase activity in the blue crab, Callinectes ornatus, acclimated to a dilute medium (21‰ salinity). This is the first kinetic demonstration of competition between spermine and spermidine for the cation sites of a crustacean Na⁺,K⁺-ATPase. Polyamine inhibition is enhanced at low cation concentrations: spermidine almost completely inhibited total ATPase activity, while spermine inhibition attained 58%; putrescine had a negligible effect on Na⁺,K⁺-ATPase activity. Spermine and spermidine affected both V and K for ATP hydrolysis but did not affect ouabain-insensitive ATPase activity. ATP hydrolysis in the absence of spermine and spermidine obeyed Michaelis–Menten behavior, in contrast to the cooperative kinetics seen for both polyamines. Modulation of V and K by K⁺, Na⁺, NH₄ ⁺ and Mg²⁺ varied considerably in the presence of spermine and spermidine. These findings suggest that polyamine inhibition of Na⁺,K⁺-ATPase activity may be of physiological relevance to crustaceans that occupy habitats of variable salinity.     

Magnesium ion effect on the helix-coil transition of DNA

The effect of magnesium ions on the parameters of the DNA helix-coil transition has been studied for the concentration range 10^?6–10^?1M at the ionic strengths of 10^?3M Na⁺. Special attention has been given to the region of low ion concentrations and to the effect of polyvalent metallic impurities present in DNA. It has been shown that binding with Mg⁺⁺ increases the DNA stability, the effect being observed mainly in the concentration range 10^?6–10^?4M. At[Mg⁺⁺]>10^?2M the thermal stability of DNA starts to decrease. The melting range extends to concentrations ～10^?5M and then decreases to 7–8°C at the ion content of 10^?3M. Asymmetry of the melting curves is observed at low ionic strengths ([Na⁺] = 10^?3M) and [Mg⁺⁺] ? 10^?5M. The results, analyzed in terms of the statistical thermodynamic theory of double-stranded homopolymers melting in the presence of ligands, suggest that the effects observed might be due to the ion redistribution from denatured to native DNA. An experimental DNA–Mg⁺⁺ phase diagram has been obtained which is in good agreement with the theory. It has been shown that thermal denaturation of the system may be an efficient method for determining the ion-binding constants for both native and denatured DNA.     

Viscosity and sedimentation study of sonicated DNA–proflavine complexes

The intrinsic viscosity of sonicated calf thymus DNA (molecular weight 4–5 × 10⁵) increases and the sedimentation constant decreases, with increasing binding of proflavine at 0. 2 ionic strength and at 25°C. The measurements correspond to a linear increase in length of the almost rodlike DNA molecules with the amount of proflavine bound; independent calculations from viscosity and sedimentation measurements yield almost identical results. Over the range of r (moles of proflavine bound per moles of nucleotides) equal to zero to r = 0.13, the length increases by about 20%. This extension is compatible with the intercalation hypothesis proposed by Lerman. Density increments at various values of r, at constant chemical potential of diffusible solutes, were determined. It was also found that, in addition to the known isosbestic point of DNA-proflavine complexes at 455.5 mμ, an additional isosbestic point exists at 225.5 mμ; this proved extremely useful for the evaluation of binding studies.     

The binding of Mg++ ions to polyadenylate, polyuridylate, and their complexes

The binding of Mg ⁺⁺ to polyadenylate (poly A), Polyuridylate(poly U), and their complexes, poly (A + U) and poly (A + 2U), was studied by means of a technique in which the dye eriochrome black T is used to measure the concentration of free Mg^?. The apparent binding constant K_X = [MgN]/[Mg⁺⁺][N], N = site for Mg⁺⁺ binding (the phosphate group of the nucleotide), was found to decrease rapidly as the extent of binding increased and, at low extents of binding, as the concentration of Na^? increased in poly A, poly (A + U), and poly (A + 2U), and somewhat less so in poly U. K_x is generally in the range 10⁴ > K_X > 10². The cause of these dependences is apparently, primarily, the displacement of Na⁺ by Mg⁺⁺ in poly U and poly (A + U) on the basis of the similarity of extents of displacement measured in this work and those measured potentiometrically. was calculated and was found to approach zero as the concentration of Na⁺ increased. In poly U, poly (A + U), and poly (A + 2U) at low ΔH′ v.H. > 0, about + 2 kcal/“mole.” In poly A, also at low salt, ΔH′ v.H. ≈ ?4 kcal/“mol” for the initial binding of Mg⁺⁺, and increases to +2 kcal/“mol” at saturation. This enthalpic variation probably accounts for the anticooperativity in the binding of Mg⁺⁺ not ascribable to the displacement of Na⁺⁺.     

Interaction of magnesium ions with poly A and poly U

The binding of Mg⁺⁺ to poly A and poly U has been measured quantitatively by using the metallochromic indicator calmagite. The method is described in detail. It is shown that there is electrostatic interaction between the binding sites, viz., the phosphate groups, and the intrinsic association constant, for the specific binding can be determined. After extrapolation to zero ionic strength we find that, for the binding of Mg⁺⁺ to poly A, k_int = 4 × 10⁴ and for that, to poly U, k_int = 3 × 10⁴. The intrinsic enthalpy of association is negative. The effect of Mg⁺⁺ on the secondary structure of poly A and poly U has been studied by measuring the ultraviolet absorbance, optical rotatory dispersion and viscosity as a function of the amount of added Mg⁺⁺ ions. It was found that Mg⁺⁺ promotes the formation of a more ordered secondary structure by neutralizing or screening the negative charges. It is concluded from the absorbance measurements that for poly A at pH ? 7 and for poly U at pH >xs 9 this ordering involves stacking of the bases. Likewise, in solutions of UDP with a pH around 10, base stacking occurs on addition of Mg⁺⁺.     

Thermodynamic and kinetic parameters of ion condensation to polynucleotides: Outer sphere complex formed by Mg++ ions

The coupling of ion binding to the single strand helix—coil transition in poly (A) and poly(C) is used to obtain information about both processes by ion titration and field-jump relaxation methods. Characterisation of the field-jump relaxation in poly(C) at various concentrations of monovalent ions leads to the evaluation of a stability constant K = 71 M^?1 for the ion binding to the polymer. The rate constant of helix formation is found to be 1.3 × 10⁷ s^?1, whereas the dissociation rate is 1.0 × 10⁶ s^?1. Similar data are presented for poly (A) and poly (dA).The interaction of Mg⁺⁺ and Ca⁺⁺ with poly (A) and poly (C) is measured by a titration method using the polymer absorbance for the indication of binding. The data can be represented by a model with independent binding “sites”. The stability constants increase with decreasing salt concentration from 2.7 × 10⁴ M^?1 at medium ionic strengths up to 2.7 × 10⁷ M^?1 at low ionic strength. The number of ions bound per nucleotide residue is in the range 0.2 to 0.3. Relaxation time constants associated with Mg⁺⁺ binding are characterised over a broad range of Mg⁺⁺ concentrations from 5 μM to 500 μM. The observed concentration dependence supports the conclusion on the number of binding places inferred from equilibrium titrations. The rate of Mg⁺⁺ and Ca⁺⁺ association to the polymer is close to the limit of diffusion control (k_R = 1 × 10¹⁰ to 2 × 10¹⁰ M^?1 s^?1). This high rate demonstrates that Mg⁺⁺ and Ca⁺⁺ ions do not form inner-sphere complexes with the polynucleotides. Apparently the distance between two adjacent phosphates is too large for a simultaneous site binding of Mg⁺⁺ or Ca⁺⁺, and inner sphere complexation at a single phosphate seems to be too weak. The data support the view that the ions like Mg⁺⁺ and Ca⁺⁺ surround the polynucleotides in the form of a mobile ion cloud without site binding.     

Sensitivity of mitochondrial Mg++ flux to reagents which affect K+ flux

Effects on Mg⁺⁺ transport in rat liver mitochondria of three reagents earlier shown to affect mitochondrial K⁺ transport have been examined. The sulfhydryl reactive reagent phenylarsine oxide, which activates K⁺ flux into respiring mitochondria, also stimulates Mg⁺⁺ influx. The K⁺ analog Ba⁺⁺, when taken up into the mitochondrial matrix, inhibits influx of both K⁺ and Mg⁺⁺. The effect on Mg⁺⁺ influx is seen only if Mg⁺⁺, which blocks Ba⁺⁺ accumulation, is added after a preincubation with Ba⁺⁺. Thus the inhibition of Mg⁺⁺ influx appears to require interaction of Ba⁺⁺ at the matrix side of the inner mitochondrial membrane. Added Ba⁺⁺ also diminishes observed rates of Mg⁺⁺ efflux but not K⁺ efflux. This difference may relate to a higher concentration of Ba⁺⁺ remaining in the medium in the presence of Mg⁺⁺ under the conditions of our experiments. Pretreatment of mitochondria with dicyclohexylcarbodiimide (DCCD), under conditions which result in an increase in the apparentK _m for K⁺ of the K⁺ influx mechanism, results in inhibition of Mg⁺⁺ influx from media containing approximately 0.2 mM Mg⁺⁺. The inhibitory effect of DCCD on Mg⁺⁺ influx is not seen at higher external Mg⁺⁺ (0.8 mM). This dependence on cation concentration is similar to the dependence on K⁺ concentration of the inhibitory effect of DCCD on K⁺ influx. Although mitochondrial Mg⁺⁺ and K⁺ transport mechanisms exhibit similar reagent sensitivities, whether Mg⁺⁺ and K⁺ share common transport catalysts remains to be established.Abbreviations used: DCCD, dicyclohexylcarbodiimide; PheAsO, phenylarsine oxide.     

Polyamine als Stimulatoren von Genaktivitäten Eine autoradiographische Studie an Riesenchromosomen von Chironomus thummi

Zusammenfassung Inaktive Riesenchromosomen der Speicheldrüsen ausgewachsener Chironomus thummi-Larven zeigten nach 60 min Inkubation mit 0,05 N Putrescin-, Spermidin-oder Spermin-Lösung erneute Syntheseaktivitäten, die sich im Einbau von radioaktiv markiertem Uridin autoradiographisch zu erkennen geben. Diese Beobachtung wird im Zusammenhang mit der bekannten genaktivierenden Wirkung von Mg⁺⁺ diskutiert. Auf der Basis zahlreicher biochemisch analoger Wirkungen von Polyaminen und Mg⁺⁺ wird ein ursprünglich für die genaktivierende Wirkung von Mg⁺⁺ postuliertes Modell auf Polyamine übertragen.

---

Stimulation of gene activities by polyamines an autoradiographic study with giant chromosomes of Chironomus thummi

Summary Inactive salivary gland giant chromosomes of full grown Chironomus thummi larvae showed after 60 min of incubation in 0.05 N putrescine, spermidine or spermine solutions autoradiographically demonstrable incorporation of radioactiv labelled uridine caused by a reactivation of RNA synthesis. This observations is discussed in relation to the known gene activating effect of Mg⁺⁺, and, since polyamines and Mg⁺⁺ have numerous effects in common, it is suggested that polyamines affect gene activity in a way similar to Mg⁺⁺.

     

The effects of cations and diamines on the viscosity of T2 DNA

The viscosity of the DNA of T2 bacteriophage has been studied with the aim of understanding the folding of the DNA in the phage head. Thus the results of previous workers have been extended into a range of concentrations and mixtures of cations and polyamines simulating the conditions obtaining in actual phage heads. Difficulties in obtaining reproducible results with DNA in 0.001M NaCl have been attributed to the presence of traces of protein, perhaps nulease(s), that can be eliminated by hot-phenol extraction of the DNA. The results have been negative in the sense that the minimum viscosity obtained (120–140 dl/g) indicates a molecule that is far from tightly folded and in that viscosities in this range can be obtained with a simple mixture of Na⁺ and Mg⁺⁺. No specificity of the diamines putrescine and spermidine is seen; they are simply surrogate cations.     

Characterization of the binding of neomycin/paromomycin sulfate with DNA using acridine orange as fluorescence probe and molecular docking technique

The binding of neomycin sulfate (NS)/paromomycin sulfate (PS) with DNA was investigated by fluorescence quenching using acridine orange (AO) as a fluorescence probe. Fluorescence lifetime, FT-IR, circular dichroism (CD), relative viscosity, ionic strength, DNA melting temperature, and molecular docking were performed to explore the binding mechanism. The binding constant of NS/PS and DNA was 6.70 × 10³/1.44 × 10³ L mol^?1 at 291 K. The values of ΔH^θ, ΔS^θ, and ΔG^θ suggested that van der Waals force or hydrogen bond might be the main binding force between NS/PS and DNA. The results of Stern–Volmer plots and fluorescence lifetime measurements all revealed that NS/PS quenching the fluorescence of DNA–AO was static in nature. FT-IR indicated that the interaction between DNA and NS/PS did occur. The relative viscosity and melting temperature of DNA were almost unchanged when NS/PS was introduced to the solution. The fluorescence intensity of NS/PS–DNA–AO was decreased with the increase in the ionic strength. For CD spectra of DNA, the intensity of positive band at nearly 275 nm was decreased and that of negative band at nearly 245 nm was increased with the increase in the concentration of NS/PS. The binding constant of NS/PS with double-stranded DNA (dsDNA) was larger than that of NS/PS with single-stranded DNA (ssDNA). From these studies, the binding mode of NS/PS with DNA was evaluated to be groove binding. The results of molecular docking further indicated that NS/PS could enter into the minor groove in the A–T rich region of DNA.     

Interactions des ions métalliques avec le DNA. IV. Fixation de i'ion cuivrique sur le DNA

The binding of cupric ion (Cu⁺⁺) to DNA was followed by spectrophotometry, melting profiles, and hydrodynamic techniques, in 0. 1M NaClO₄ and at pH 5. 6. A small amount of Cu⁺⁺ is bound specifically to bases (about 1 Cu⁺⁺ per 20 nucleotides), in agreement with polarographic and EPR data. A preferential stabilization of G–C pairs and only a slight increase of the flexibility of the molecule were observed. In 5 × 10^?3M NaClO₄, a higher number of nonhomogeneous binding sites is found by spectrophotometry. It is concluded that at least two types of sites are available for Cu⁺⁺. The first one, where Cu⁺⁺ is chelating N₇ of purines to phosphate, is observed only at low ionic strength and destabilizes the double helix. The second exists mainly at 0, 1M or higher ionic strength. All the sites are identical and could be attributed to two successive guanine residues in the same strand. Similar behavior was found for other divalent cations, e. g., Fe⁺⁺, Mn⁺⁺, and Co⁺⁺.     

Binding of cations by microsomes from rabbit skeletal muscle

Fragmented sarcoplasmic reticulum and transverse tubular system, as isolated in the microsomal fraction from rabbit skeletal muscle, bind H⁺, Na⁺, K⁺, Ca⁺⁺, Mg⁺⁺, and Zn⁺⁺. The binding depends on a cation exchange type of interaction between these cations and the chemical components of the membranous systems of the muscle cell. The monovalent and divalent cations exchange quantitatively for each other at the binding sites on an equivalent basis. Scatchard plots of the H⁺ binding data indicate that the binding groups can be resolved into two major components in terms of their pK values. Component 1 has a pK value of 6.6 and a capacity for H⁺ binding of 2.2 meq/g N . The second component has a much higher H⁺ binding capacity (7–8 meq/g N ), but its pK value, 3.4, is non-physiological. The binding of cations other than H⁺ at a neutral pH occurs at the binding sites making up component 1. The order of affinity of the cations for the microsome binding sites is H⁺ » Zn⁺⁺ > Ca⁺⁺ > Mg⁺⁺ » Na⁺ = K⁺ as reflected by the apparent respective pK_M values: 6.6, 5.2, 4.7, 4.2, 1.3, 1.3. Caffeine, which causes contracture and potentiates the twitch of skeletal muscle, does not interfere with the binding of Ca⁺⁺ by the microsomes at neutral pH.     

Binding of ethidium to bacteriophage T7 and T7 deletion mutants

Equilibrium binding of ethidium, quantitated by fluorescence enhancement, to DNA packaged in bacteriophage T7 and T7 deletion mutants has been compared with the binding of this dye to DNA released from its capsid (free DNA). During achievement of apparent equilibrium binding, no change in bacteriophage T7 structure occurred, by the criterion of agarose gel electrophoresis. However, excessive incubation with ethidium bromide caused detectable changes in bacteriophage structure, a possible explanation of disagreements in similar studies previously performed with T-even bacteriophages. Scatchard plots for packaged DNA had a curvature greater than the previously demonstrated [Bresloff, J. L. & Crothers, D. M. (1981) Biochemistry 20 , 3547–3553] curvature for free DNA. By treating plots for packaged DNA as though they were biphasic, it was found that binding to most sites occurred with an apparent association constant (K_ap) 3.3–4.3 times lower than the K_ap of free DNA. The number of these sites increased significantly as the density of packaged DNA was decreased by use of the deletion mutants. Values of ΔH° for these sites were negative and equal to the ΔH° for free DNA; values of ΔS° were positive and about half the ΔS° for free DNA. A second class of sites, roughly 1.2% of the total, had a significantly higher K_ap and more negative ΔH° than those of the majority of sites.     

The effect of neutral salts on the conformational transition of poly-alpha-amino acids. I. Potentiometric titration and optical rotatory dispersion studies

The effect of salts on the coil-to-helix transition of poly-α-amino acids was investigated by optical rotatory dispersion and potentiometric titration techniques. Both charge-dependent and charge-independent contributions to the free energy were considered. The free energy of formation ΔF° of the uncharged α-helix from the uncharged random coil for poly-L -glutamic acid (PGA) decreases very rapidly in the limit of zero added salt concentration. This effect probably depends on the uncertainty affecting the choice of the extrapolation of the apparent pK for the random coil at low ionic strength. Above 0.1 M salt, where the free energy determination becomes meaningful, the anions and cations investigated do not affect the value of ΔF°, with the exception of Li⁺. Our data support the point of view that this cation binds to the peptide group. A class of salts produces an increase of the helical content of poly-L -ornithine (PO) both at low and high degree of ionization. This effect appears to be anion dependent. It is shown that (1) no change of ΔF° is involved; (2) recent theories of polyelectrolyte solutions cannot account for our results. We suggest that a true site binding of the anions to the charged amino groups occurs. The role of electrostatic binding in determining the conformational stability of proteins in the presence of some anions is stressed, and a general treatment for the electrostatic binding equilibria is outlined.     

Equilibrium studies on neutral red-DNA binding.

Spectrophotometric binding studies were undertaken on the interaction of neutral red with native and heat-denatured, sonicated, calf thymus DNA in a 0.2M ionic strength buffer containing Tris–sodium acetate–potassium chloride at 25°C. The pK_A of neutral red was found to be 6.81. At pH 5 the binding of protonated neutral red was complicated even at low concentration ratios of dye to DNA. In the pH range 7.5–8.5 the tight binding process could be studied and it was found that both protonated and free base species of neutral red significantly bind with DNA having association constants (in terms of polynucleotide phosphate) of 5.99 × 10³ M^?1 and 0.136 × 10³ M^?1, respectively, for native DNA and 7.48 × 10³ M^?1 and 0.938 × 10³ M^?1, respectively, for denatured DNA. The pK_A value of the neutral red–DNA complexes were 8.46 for native DNA and 7.72 for denatured DNA. These results are discussed in terms of possible binding mechanisms.     

Extraction of an actomyosin-like pootein from amoebae of Dictyostelium discoideum

An actomyosin-like protein has been extracted from amoebae of Dictyostelium discoideum, V-12. The purified protein exhibited a reversible change in viscosity upon addition of ATP, indicating an ATP sensitivity of 75–85% and a specific viscosity of 0.1. At low ionic strength in the presence of Mg⁺⁺ and ATP the amoeba protein displayed the phenomenon of superprecipitation. The protein extract was found to be an adenosinetriphosphatase (ATP'ase) hydrolyzing ATP to ADP and inorganic phosphate. Both Mg⁺⁺ and Ca⁺⁺ at low ionic strength accelerated the ATP ase activity whereas at high ionic strength only Ca⁺⁺ stimulated ATP hydrolysis. The ATP'ase activity was inhibited by ethylene-diamine-tetraacetic-acid, Mersayl and p-chloromercuribenzoate. The physico-chemical and enzymatic properties of the extracted amoeba protein are qualitatively comparable to those of muscle actomyosin, and very similar in quantitative properties to smooth muscle actomyosin and the actomyosin-like proteins of blood platelets, leucocytes and slime mold plasmodia. The significance of the presence of this actomyosin-like protein in Dictyostelium amoebae is discussed in relation to amoeboid form and movement.