Cerebrosides sulphated

It has been found that injected 35S-sulphate reaches maximal incorporation into rat brain cerebroside sulphate only 2 days after injection, and its activity remains practically constant and undergoes only very slow turnover. After 32 days the level of cerebroside sulphate activity was still 3/4 of that found on the second day after injection980. In liver, spleen and heart the maximum incorporation of 35S into cerebroside sulphate was reached after 12 hours and after 4 days the radioactivity virtually disappeared in these organs. 

However, in the kidney the maximum incorporation had been attained 24 hours after injection, and the radioactivity was still present on the 32nd day. In the brain the 35S-substance sedimented with mitochondria in the kideny with microsomes and in the liver it was found in the soluble fraction. No cerebroside sulphate has been observed in rat blood cells or plasma. 

Sulphation of lactose with pyridine-SOa complex in pyridine resulted in formation of the galactose 6-sulphate. Other oligosaccharides were alsostudied. I he glycolipids, seminolipid and cerebroside sulphate, which are sulphated on the 3-position of galactose, were not oxidized by galactose oxidase. 

L-Tyrosine O-sulphate has been shown to be hydrolysed by pure human arylsulphatase A at 5% of the rate for nitrocatechol sulphate, but at a comparable rate to that for cerebroside sulphate." The reaction value 0.35 mM for L-tyrosine 0-sulphate) was optimal at pH 5.3-5.5 and displayed zero order kinetics with time and enzyme concentration. 

In addition to cerebroside sulphates (Section 2.4) and diacylsulphoquinovosylglycerol (Section 2.5), various other sulphur-containing lipids have been reported. These include alkyl sulphates (Mayers etal., 1969) in micro-organisms. 

Fluharty et aL (1974) have reported the isolation of S-labelled cerebroside sulphate. They injected the brains of young rats intracerebrally with [ S]-sulphate and sacrificed the animals 3 days later. Total lipids were extracted and glycerolipids destroyed by alkaline hydrolysis. TEAE-cellulose column chromatography was then used to purify the [ Sjsulphatide. Jatzkewitz and Nowoczek (1967) have synthesized o-galactose 3-sulphate and shown it to be identical to the galactose sulphate present in brain sulphatides. 

Fig. 140. Thin-layer chromatogram of sphingolipids [210]. Adsorbent silica gel G solvent chloroform-methanol-water (60 + 35 + 8) time of run 2 h spray reagents diphenylamine solution and Dragendorff reagent for detection of sphingomyelin amounts 50—100 jxg of each 2 sphingolipid mixture from beef brain 2 non-esterified cerebrosides 3 cerebroside sulphates 4 sphingomyelin 5 gang-...

It has been shown that the lipid composition of the brain of mentally diseased adults is often similar to that of normal infants much more sulphatide is found in cases of metachromatic leucodystrophy [69, 72, 177] large amounts of one ganglioside appear in Tay-Sachs disease [96, 177] and a greatly increased sphingomyelin content of the brain is found with the Niemann-Pick disease [164, 177]. The cerebroside sulphate esters of the brain of normal persons and of those suffering from metachromatic leucodystrophy have been isolated by TLC and shown to be identical [69]. 

The distribution of soluble arylsulphatases in human tissues has been examined via ion-exchange chromatography. All tissues contained arylsulphatase A and B. In addition, brain singularly contained significant quantities of a minor anionic form (Bm) only trace amounts of Bm were found in liver, kidney, testis, and placenta. Arylsulphatases B and Bm had equal activity towards methylumbelliferyl sulphate, nitrocatechol sulphate, and UDP-2-acetamido-2-deoxy-D-galactose 4-sulphate, but both forms were inactive toward the arylsulphatase A substrates cerebroside sulphate and L-ascorbic acid 2-sulphate. The physicochemical properties of arylsulphatases B and Bm differed with respect to thermal stability, ionicity, and behaviour on polyacrylamide gel electrophoresis and isoelectric focusing. 

Acidic forms of human cerebroside sulphatase have been partially purified from invertebrates Tethya aurantium (Porifera), Patella vulgata (mollusca), Maja squinado (Arthropoda), Martlasterias glacialis (Echinodermata), and Microcosmus sulcatus (Tunicata). Enzyme preparations thus obtained cleaved cerebroside sulphates only in the presence of either specific detergents, e.g. taurodeoxycholate, or an activator protein isolated from human liver. On a molar basis, less of the activator protein was required to achieve the same activation as detergent. 

Arylsulphatase A from human liver and kidney has been shown to possess endogenous cerebroside sulphatase activity, and both activities appear to be manifest by a single active site. The arylsulphatase was inhibited by cerebroside sulphate in a way that depends on the ionic strength of the solvent. Differences in the two types of reaction catalysed were ascribed to differences in the physicochemical states of the substrates. 

The requirement for bile salts and manganese chloride, and the pH optimum and Km value of a purified human arylsulphatase A have been reported. Aryl sulphate and 2-0-acyl-l-0-alkyl-3-0-()3-D-galactopyranosyl 3-sulphate)glycerol were hydrolysed by the enzyme under identical conditions. The glycolipid was not hydrolysed by arylsulphatase B and appears to be the physiological substrate for arylsulphatase A. Human arylsulphatase B was not active towards cerebroside sulphate. 

An assay for cerebroside sulphatase is based on t.l.c. and the use of tritium-labelled cerebroside sulphate as a substrate. The cerebroside sulphatase activity of the arylsulphatase A in human liver and kidney was investigated using this assay. Kinetic and other evidence suggested that cerebroside sulphatase also possesses arylsulphatase activity, although both activities are manifest by the same active site. Pure human cerebroside sulphatase has been shown to require the presence of bile salts and to be stimulated by manganese chloride. Cerebroside sulphatases isolated from a number of invertebrates are composed of multiple forms that were separated by isoelectric focusing. These enzymes have activities comparable to those from vertebrates, and they also exhibit arylsulphatase activity. Kinetic data (pH optima and Km values etc.) were reported for the enzymes, which were inhibited by nitrocatechol sulphate. 

Sulphatases.—Desialylization of sulphatase A from ox liver with neuraminidase slightly modified the charge on the molecule, but did not alter any other properties. The modified enzyme was able to hydrolyse cerebroside sulphate. 

Cerebroside sulphate labelled with tritium has been used as a substrate in an assay for cerebroside sulphatase activities using t.l.c. ... 

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