TRANSAMIDINASE OF HOG KIDNEY. I. PURIFICATION AND PROPERTIES.

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Fructose 1,6-Diphosphatase from Rabbit Liver XII. EFFECT OF SUBSTRATE AND ADENOSINE MONOPHOSPHATE ON THE IONIZATION OF THE TYROSYL RESIDUES

Abstract The effects of substrate and AMP on the state of ionization of tyrosyl residues of fructose 1,6-diphosphatase have been investigated. In the native enzyme, 5 to 6 tyrosyl residues were titrated with a pK of about 8.4, and 7 to 8 residues were titrated with a pK of 9 to 9.2. The remaining tyrosyl residues were not titrated below pH 10. In the presence of 10-5 m fructose 1,6-diphosphate, when 4 moles of substrate are bound per mole of enzyme, approximately 4 of the low pK tyrosyl residues are shifted from pK 8.4 to pK 9.7. However, all of the pK values are modified, and under these conditions 2 tyrosyl residues show a pK of 8.7, and 10 or 11 residues show a pK of 9.7. At a high concentration of fructose 1,6-diphosphate, 10-3 m, all 12 to 13 of the tyrosyl residues are titrated with a pK of 9.8. The results can be correlated with previous studies on the acetylation of fructose 1,6-diphosphatase. The tyrosyl residues with low pK are more reactive and are associated with loss of sensitivity to AMP. However, after acetylation, only 10 tyrosyl residues are titrated with a pK of 9.3 to 9.4, and the pK is shifted to 9.7 when the substrate is bound. A method of analyzing the titration data is presented which permits simultaneous evaluation of number of residues and pK value of each titrated group. This has revealed changes in tyrosyl residues that would otherwise have been overlooked

Studies on the mechanism of action of the gluconate 6-phosphate dehydrogenase. The presence of a cysteine residue in the active center.

Abstract The role of the sulfhydryl groups in the gluconate 6-phosphate dehydrogenase has been studied. Nine cysteine residues have been detected by spectrophotometric titration with p-hydroxymercuribenzoate, but the reaction of only 3 of them is sufficient to inactivate the enzyme completely. Chlorodinitrobenzene acts even more specifically. Incorporation of 1.6 dinitrobenzene residues leads to enzyme inactivation. The amino acid residues involved in dinitrophenylation have been identified as cysteine. Gluconate 6-phosphate completely protects the enzyme against inactivation by chlorodinitrobenzene

THE NATURE OF THE AMINO ACID RESIDUES INVOLVED IN THE INACTIVATION OF GLUCONATE 6-PHOSPHATE DEHYDROGENASE BY IODOACETATE.

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The actin gelling activity of chicken gizzard α-actinin at physiological temperature is triggered by water sequestration

AbstractAt 37°C, in the presence of 6% (wv) polyethylene glycol 6000, 30 nM α-actinin from chicken gizzard induces the gelation of 12 μM actin. Static measurement shows that the addition of 30 nM α-actinin increases the rigidity of the system from 23.5 to 54 dynescm2. According to the theory of osmoelastic coupling, also large additives, such as the proteins of the cell sap, are able to cause an osmotic stress equivalent to that caused by polyethylene glycol. We thus conclude that, in vivo, α-actinin acts as an actin gelling protein

Conformational states of rabbit liver fructose 1,6-diphosphatase.

Abstract An analysis of the conformational states of fructose 1,6diphosphatase by the circular dichroism technique has been performed. It has been shown that the enzyme presents a strong rigidity of its over-all conformation. The addition of either the substrate or the allosteric inhibitor, AMP, or changes in the pH of the medium, do not produce significant modifications of the secondary and tertiary structure of the protein. The same situation holds even in 0.05 m sodium dodecyl sulfate, a condition in which the quaternary structure of the enzyme undergoes a profound modification. Small conformational changes have been detected, however, after the addition of the substrate and of AMP. These changes appear to be restricted to a limited number of tyrosyl and, at maximum, to a few vicinal residues. It is concluded that the catalytic and regulatory functioning of the enzyme do not require substantial alterations in the secondary and tertiary structure of the protein, but are related to changes in the ionization of amino acid residues and to subtle variations of the quaternary structure

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

AbstractIn 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

Skeletal muscle contraction. The thorough definition of the contractile event requires both load acceleration and load mass to be known

<p>Abstract</p> <p>Background</p> <p>The scope of this work is to show that the correct and complete definition of the system of muscle contraction requires the knowledge of both the mass and the acceleration of the load.</p> <p>Results</p> <p>The aim is achieved by making use of a model of muscle contraction that operates into two phases. The first phase considers the effects of the power stroke in the absence of any hindrance. In the second phase viscous hindrance is introduced to match the experimental speed and yield of the contraction. It is shown that, at constant force of the load, changing load acceleration changes the time course of the pre-steady state of myofibril contraction. The decrease of the acceleration of the load from 9.8 m.s^-2to 1 m.s^-2increases the time length of the pre-steady state of the contraction from a few microseconds to many hundreds of microseconds and decreases the stiffness of the active fibre.</p> <p>Conclusions</p> <p>We urge that in the study of muscle contraction both the mass and the acceleration of the load are specified.</p