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Biomedical subjects

A B Roy

Publications and source records attributed to A B Roy.

At least 55 records · Page 3Linked to original sources
PubMed1980

The sulphatase of ox liver. XXIII. The nature of substrate-modified sulphatase A.

An improved method is described for the preparation of milligram quantities of substrate-modified sulphatase A. The latter has the same molecular weight and the same ability to form a tetramer as has native sulphatase A. It has been shown that the modified enzyme prepared with nitrocatechol [35S]sulphate as substrate contains 1 mol 35SO24- per mol enzyme and that any treatment which causes reversion of the modified enzyme to native enzyme is accompanied by the loss of the bound SO24-. Dialysis of the 35S-modified enzyme against a solution containing SO24- causes a loss of 35SO24- with no change in the amount of modified enzyme in the preparation. It has been shown that the activation of the substrate-modified enzyme by SO24- does not lead to the formation of a third stable form of sulphatase A.

Animals↗
PubMed1979

The sulphatase of ox liver. XXII. Further observations on the cerebroside sulphatase activity of sulphatase A.

Further studies have been made of the cerebroside sulphatase activity of the sulphatase A (aryl-sulphate sulphohydrolase, EC 3.1.6.1) of ox liver. It is concluded that a cerebroside sulphate-modified form of the enzyme is not produced and that the kinetics of the reaction can be explained by the utilisation of the substrate and accumulation of (SO4)2-. The hypothesis is advanced that this difference between the cerebroside sulphatase and arylsulphatase activities arises from non-polar binding of the cerebroside to the enzyme. Possible reasons for the differences between these results and those of other (Stinshoff, K. and Jatzkewitz, H. (1975) Biochim. Biophys. Acta 377, 126-138) are considered.

Animals↗
PubMed1979

Sulphatase A: an arylsulphatase and a glycosulphatase.

Sulphatase A was first described as an arylsulphatase but was subsequently shown to have cerebroside sulphatase activity, bydrolysing galactose 3-sulphate residues in certain lipids. A characteristic feature of the arylsulphatase activity is the substrate-induced inactivation of the enzyme which occurs during the catalytic reaction. Present views of the course of this modification are considered. A similar modification of the enzyme does not occur during the cerebroside sulphatase reaction and the fall in velocity noted during most such assays can be explained by disappearance of substrate and accumulation of sulphate. Possible reasons for the difference between the two types of activity are considered. Sulphatase A also hydrolyses hexose sulphates at rates comparable to those of aryl sulphates. The specificity of sulphatase A towards its natural substrates, sulpholipids, is considered in the light of these findings.

Animals↗
PubMed1978

Purification and properties of the ATP sulphurylase of rat liver.

ATP sulphurylase (ATP:sulphate adenylyltransferase, EC 2.7.7.4) as been purified about 2500-fold from rat liver. It was free of ATPase, inorganic pyrophosphatase, adenosine phosphosulphate kinase and ADP sulphurylase activities. The enzyme was homogeneous to chromatography on Sepharose 4B and to density-gradient sedimentation; it was not homogeneous to acrylamide gel electrophoresis nor to sedimentation in the ultracentrifuge. Possible reasons for this heterogeneity are considered. The molecular weight of the enzyme is 410 000 as measured by chromatography on Sepharose 4B. The v is 0.80, suggesting that ATP sulphurylase is a lipoprotein. The enzyme activity is associated with a pigment having a lambdamax of 410 nm. Studies of the forward, reverse and ATP-PPi exchange reactions catalysed by ATP sulphurylase have shown that these are sequential bi-bi reactions, with ATP being the first substrate bound and adenosine phosphosulphate the last product released. The results are incompatible with previous suggestions that the ATP sulphurylase of rat liver catalysed a bi-bi ping-pong reaction.

Animals↗
PubMed1978

The sulphatase of ox liver. XXI: kinetic studies of the substrate-induced inactivation of sulphatase A.

The theoretical basis is given for methods of determining the apparent velocity constant, k*, for the substrate-induced inactivation of sulphatase A (aryl-sulphate sulphohydrolase, EC 3.1.6.1) and the initial velocity, vo, of the catalytic reaction. The expression is of the same form as the empirical relationships previously used but the significance of the various terms is clearly established. The method has been applied to the characterisation of the inactivation occurring during the hydrolysis of a number of substrates and it has been shown that k* varies with so in a hyperbolic relationship described by k, a velocity constant at infinite substrate concentrations and by K, a constant analogous to the Michaelis constant. Although K varies considerably for different substrates, and is consistently less than the corresponding Km, k is almost constant at 0.23 min-1. It is therefore suggested that the inactivation of the enzyme does not proceed through an enzyme . substrate complex but through the enzyme . SO2-4 complex produced during the catalytic reaction. The effects of several variables on these parameters are described.

Animals↗
PubMed1977

Micelles of cerebroside sulphate.

The critical micelle concentration of cerebroside sulphate in water is 0-01 mM: it increases with increasing concentrations of buffer to 0-07 mM in 0-1 M sodium acetate and formate buffers, pH 5-6 and 4-5 respectively. The partial specific volume of the micelles is about 0-94. The behaviour of the micelles in the ultracentrifuge and on Sephadex G-200 shows them to be grossly heterogeneous with respect to size. In 0-1 M buffer s20,w is about 26 S; in water or 0-01 M buffer smaller micelles with an s20,w of about 6 S are also present. In 0-01 M formate, pH 4-5, the smallest species detectable by equilibrium ultracentrifugation had a micellar weight of about 180,000 corresponding to an aggregation number of about 180. Much larger aggregates were also present. It is suggested that the smallest micelles are the substrate for sulphatase A when this is acting as a cerebroside sulphatase in buffers of low ionic strength.

Animals↗
PubMed1976

The sulphatase of ox liver. XX. The preparation of sulphatases B1alpha and B1beta.

Sulphatases B1alpha and B1beta (EC 3.1.6.1) have been prepared as apparently homogeneous proteins by chromatography on ConA-Sepharose. Both have a mol. wt. of 56 000, and E1%280nm of 17 and a turnover number of 8600 min-1 with nitrocatechol sulphate as substrate. Their amino acid compositions are identical: like sulphatase A, the sulphatases B are rich in proline and yield glucosamine on hydrolysis. They are not altered by treatment with neuraminidase. Both fractions show strong UDP-N-acetylgalactosamine 4-sulphatase activity, weak iduronate sulphatase activity, but no significant heparan N-sulphatase activity. It is suggested that the physiological activity of sulphatase B is that of the N-acetylgalactosamine 4-sulphatase which is lacking in the Maroteaux-Lamy Syndrome.

Amino Acids↗
PubMed1976

The sulphatase of ox liver. XIX. On the nature of the polymeric forms of sulphatase A present in dilute solutions.

Weight-average elution volumes of sulphatase A (an arylsulphate sulphohydrolase, EC 3.1.6.1) from Sephadex G-200 have been determined as functions of protein concentration, pH, ionic strength and temperature. The results are used to calculate the apparent association equilibrium constants for tetramer formation and the associated standard-state thermodynamic parameters. While the apparent association constant decreased from 10(28) to 10(21) M-3 on increasing the pH from 4.5 to 5.6 at ionic strength 0.1, at any particular pH value studied it was relatively insensitive to temperature variation so that deltaH is close to zero and tetramer formation in solution is associated with a positive entropy change. At pH 5.0, increasing the ionic strength from 0.1 to 2 decreased the association constant by a factor of 100. Methylumbelliferone sulphate has no effect on the association of sulphatase A. The equilibrium results are used to define the degree of association of sulphatase A likely to encountered in experiments designed to elucidate its kinetic properties. In the liver lysosome, the tetramer is probably the dominant species. The monomer and tetramer of sulphatase A have similar, or identical, specific activities with nitrocatechol sulphate and 4-methylumbelliferone sulphate as substrates. With nitrocatechol sulphate, sulphatase A shows Michaelis kinetics under conditions where the monomer is the dominant species and non-Michaelis kinetics where the tetramer is dominant. There is apparently a negative cooperativity between the monomer units in the tetramer. In 2 mM sodium taurodeoxycholate and 0.035 M MnCl2, but not in 0.1 M NaCl, the tetramer shows Michaelis kinetics. This is not due to dissociation of the tetramer. The critical micellar concentration of sodium taurodeoxycholate is about 0.8 mM in both 0.1 M NaCl and 0.035 M McCl2 but the aggregation number is greater in the latter.

Animals↗
PubMed1975

L-ascorbic acid 2-sulphate. A substrate for mammalian arylsulphatases.

Ascorbic acid 2-sulphate has a stability in acid comparable to that of phenyl sulphate and is rather more acid-labile than simple carbohydrate sulphates. At its optimum pH of 4.8 sulphatase A(aryl-sulphate sulphohydrolase EC 3.1.6.1.) hydrolyses ascorbic acid sulphate with a specific activity of 90 mumol/mg per min (150 mumol/mg per min with nitrocatechol sulphate at pH 5.6). At pH 4.8 the kinetics are non-Michaelis. At pH 5.6 Michaelis kinetics are obeyed and Km 12 21 mM ascorbic acid 2-sulphate. K2SO4 is a competitive inhibitor with a Ki of 0.2 and 0.6 mM at pH 4.8 and 5.6, respectively. Sulphatase A is converted into a substrate-modified form during its hydrolysis of ascorbic acid sulphate. Sulphatase B also hydrolyses ascorbic acid 2-sulphate. At pH 4.8 and in the presence of 0.15 M NaCl the specific activity is 0.92 mumol/mg per min (90 mumol/mg per min for nitrocatechol sulphate at pH 5.6). In the absence of NaCl the activity is greatly decreased. Km is 8 mM. K2SO4 is a competitive inhibitor with a Ki of 0.1 mM. Ascorbic acid is not hydrolysed at a detectable rate by the arylsulphatases of the mollusc Dicathais orbita or of Aerobacter aerogenes.?

Animals↗