[115] Characterization of phosphorus compounds by acid lability

Leloir, Luis F.; Cardini, Carlos E.

doi:10.1016/s0076-6879(57)03459-x

Cited by 268 publications

(110 citation statements)

References 16 publications

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“…RNA was assayed by the orcinollmethod (Munro & Fleck, 1966) with D-ribose as standard, and amino acids by the ninhydrin reaction (Rosen, I 957) with L-leucine as standard. Samples for determination of tota1"phosphorus were charred, oxidized and hydrolysed (Leloir & Cardini, 1957), and the inorganic phosphate produced was assayed by the method of Stanton (1968).…”

Section: Methodsmentioning

confidence: 99%

Permeability of Dictyostelium discoideum towards Amino Acids and Inulin: a Possible Relationship between Initiation of Differentiation and Loss of 'Pool' Metabolites

Lee

1972

Journal of General Microbiology

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The permeability of Dictyostelium discoideum amoebae towards amino acids and inulin was studied under conditions suitable for development of aggregationcompetence and in thick cell suspensions. During the pre-aggregation period, the amoebae released large amounts of amino acids and nucleotides into the suspending medium. The uptake of glutamate and lysine by amoebae was passive, but inulin was taken up actively, probably by pinocytosis. Hence the simplest explanation for leakage of metabolites is that the amoebae have no active mechanism for retaining them. A possible relationship between initiation of differentiation and loss of pool metabolites is discussed.

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Section: Methodsmentioning

confidence: 99%

Permeability of Dictyostelium discoideum towards Amino Acids and Inulin: a Possible Relationship between Initiation of Differentiation and Loss of 'Pool' Metabolites

Lee

1972

Journal of General Microbiology

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“…Acid hydrolyis gave 98-101 % of the theoretical amount of inorganic phosphate (Leloir and Cardini 1957). and assay with crystalline lactate dehydrogenase indicated that the preparation contained less than 0·5% pyruvate.…”

Section: (B) Ghemical8mentioning

confidence: 97%

Succinic Acid Production by Rumen Bacteria. III. Enzymic Studies on the Formation of Succinate by Ruminococcus Flavefaciens

Hopgood¹,

Walker²

1969

Aust. Jnl. Of Bio. Sci.

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SummaryEnzymes involved in succinic acid production by strain C of R. ftavefacien8 were investigated in cell-free extracts. The results indicate that phosphoenolpyruvate is carboxylated by a phosphoenolpyruvate carboxykinase which uses GDP as phosphate acceptor, and that the oxaloacetate so formed is converted to succinate via a DPNH-dependent malate dehydrogenase, a fumarate hydratase, and a DPNHdependent fumarate reductase. Succinate dehydrogenase activity was also observed which differed markedly from fumarate reductase in that DPN+ was not reduced and in inhibition characteristics.There is no evidence that synthesis of ATP is associated with the conversion of oxaloacetate to succinate, or that DPNH oxidation is coupled to ATP synthesis. It is concluded that approximately 2·8 moles of ATP are formed per mole of glucose fermented by this organism.

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“…The ATPase activity was measured using the coupled enzyme system [26], except for kinetic assays in the absence of Mg2+ when it was measured using the method of Fiske and Subba-Row [27]. The assay mixture contained in a final volume of 1 ml the reagents listed in the figure legends and the following detection system: pyruvate kinase (amount yielding 7.5 pmol product/min), lactate dehydrogenase (amount yielding 18 pmol product/min), 0.25 mM NADH and 0.42 mM phosphoerzolpyruvate.…”

Section: Measurement Of Atpase Activitymentioning

confidence: 99%

Distances between functional sites of the Ca²⁺+ Mg²⁺‐ATPase from sarcoplasmic reticulum using Co²⁺ as a spectroscopic ruler

Cuenda¹,

Henao²,

Gutiérrez‐Merino³

1990

European Journal of Biochemistry

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Cobalt ion inhibits the Ca2+ + Mg2 +-ATPase activity of sealed sarcoplasmic reticulum vesicles, of solubilized membranes and of the purified enzyme. To use Co2+ appropriately as a spectroscopic ruler to map functional sites of the Ca2+ + Mg2+-ATPase, we have carried out studies to obtain the kinetic parameters needed to define the experimental conditions to conduct the fluorimetric studies.1. The apparent Ko.5 values of inhibition of this ATPase are 1.4 mM, 4.8 mM and 9.5 mM total Co2+ at pH 8.0, 7.0 and 6.0, respectively. The inhibition by Co2+ is likely to be due to free Co2+ binding to the enzyme. Millimolar Ca2 from which we obtain a distance of 1.1 -1.9 nm between Co2+ and fluorescein located at neighbouring catalytic sites. Co2+. ATP can be used as a substrate by this enzyme with V,,, of 2.4 f 0.2 pmol ATP hydrolyzed min- Co2+ partially quenches, about 10The sarcoplasmic reticulum Ca2+ + Mg2 +-ATPase uses Mg2+ . ATP as its physiological substrate and couples ATP hydrolysis to the active transport of Ca" across this membrane [l]. Binding of two Ca2+ ions to high-affinity binding sites/ATPase molecule is step that precedes Ca2+ transport to the luminal space of sarcoplasmic reticulum [l, 21. To understand better the Ca2+ transport process coupled to ATP hydrolysis requires complementary experimental information obtained by using kinetic and structural studies of this enzyme. In this regard, the location of Ca2+ and ATP binding sites relative to each other is an important issue; there is a large range of values for the distance between these sites on ATPase reported by different groups of investigators, i. e. 1 -6 nm [3 -71. Forster energy transfer between different donor/acceptor pairs of fluorescent targets selectively bound to these sites has been used to obtain these estimations. On the other hand, Xray diffraction studies [8, 91 and image processing of electron microscopy data [lo, 111 have provided evidence for an overall shape of an ATPase unit protruding about 4.0 -6.0 nm from the lipid/water interface and having a cross-sectional diameter of the cytosolic lobule of approximately 8.0-9.0 nm in its dimeric form, which is probably the predominant state in

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[115] Characterization of phosphorus compounds by acid lability

Cited by 268 publications

References 16 publications

Permeability of Dictyostelium discoideum towards Amino Acids and Inulin: a Possible Relationship between Initiation of Differentiation and Loss of 'Pool' Metabolites

Permeability of Dictyostelium discoideum towards Amino Acids and Inulin: a Possible Relationship between Initiation of Differentiation and Loss of 'Pool' Metabolites

Succinic Acid Production by Rumen Bacteria. III. Enzymic Studies on the Formation of Succinate by Ruminococcus Flavefaciens

Distances between functional sites of the Ca²⁺+ Mg²⁺‐ATPase from sarcoplasmic reticulum using Co²⁺ as a spectroscopic ruler

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[115] Characterization of phosphorus compounds by acid lability

Cited by 268 publications

References 16 publications

Permeability of Dictyostelium discoideum towards Amino Acids and Inulin: a Possible Relationship between Initiation of Differentiation and Loss of 'Pool' Metabolites

Permeability of Dictyostelium discoideum towards Amino Acids and Inulin: a Possible Relationship between Initiation of Differentiation and Loss of 'Pool' Metabolites

Succinic Acid Production by Rumen Bacteria. III. Enzymic Studies on the Formation of Succinate by Ruminococcus Flavefaciens

Distances between functional sites of the Ca2++ Mg2+‐ATPase from sarcoplasmic reticulum using Co2+ as a spectroscopic ruler

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Distances between functional sites of the Ca²⁺+ Mg²⁺‐ATPase from sarcoplasmic reticulum using Co²⁺ as a spectroscopic ruler