Patricia V. Donnelly

Collagen treated with (+)-catechin becomes resistant to the action of mammalian collagenase.

Treatment of radioactively labeled guinea-pig skin soluble collagen or calf skin collagen with the flavonoid (+)-catechin makes the collagen resistant to the action of mammalian collagenase but not to the action of bacterial collagenase. Complete resistance to the action of the mammalian enzyme may be achieved by incubating 0.6 mg of collagen (dry weight) with 0.1 mM (+)-catechin, followed by dialysis to remove the unbound flavonoid. Since incubation of the mammalian enzyme with (+)-catechin does not inhibit its activity, it is postulated that (+)-catechin binds tightly to collagen and modifies its structure sufficiently to make it resistant to enzyme degradation.

Deficiencies of Glucosamine-6-Sulfate or Galactosamine-6-Sulfate Sulfatases Are Responsible for Different Mucopolysaccharidoses

[1-3H]Galactitol-6-sulfate, N-[1-3H]acetylgalactosaminitol-6-sulfate, N-[1-3H]acetylglucosaminitol-6-sulfate, N-acetylglucosamine-6-sulfate, and 6-sulfated tetrasaccharides from chondroitin-6-sulfate have been used for the measurement of 6-sulfatase activity of extracts of normal skin fibroblasts and of fibroblasts cultured from patients with genetic mucopolysaccharidoses. With these substrates, extracts of fibroblasts derived from Morquio patients lack or have greatly reduced activities for galactitol-6-sulfate, N-acetylgalactosaminitol-6-sulfate, and 6-sulfated tetrasaccharides but have normal activity for N-acetylglucosamine-6-sulfate and its alditol; those derived from a patient with a newly discovered mucopolysaccharidosis have greatly reduced activity for N-acetylglucosamine-6-sulfate and its alditol but normal activity for galactitol-6-sulfate, N-acetylgalactosaminitol-6-sulfate, and the 6-sulfated tetrasaccharides. These findings demonstrate the existence of two different hexosamine-6-sulfate sulfatases, specific for the glucose or galactose configuration of their substrates. Their respective deficiencies, causing inability to degrade keratan sulfate and heparan sulfate in one case and keratan sulfate and chondroitin-6-sulfate in the other, are responsible for different clinical phenotypes.

Spondyloepiphyseal dysplasia, chondroitin sulfate type: A possible defect of paps — Chondroitin sulfate sulfotransferase in humans

Abstract Four patients with an unusual form of spondyloepiphyseal dysplasia excreted in the urine undersulfated chondroitin 6-sulfate (Biochem. Med. 7 , 415–423, 1973). The sera of these patients show a low activity of PAPS — chondroitin sulfate sulfotransferase, while the undersulfated chondroitin sulfate present in their urine is a much better acceptor of 35 SO 4 than standard chondroitin sulfate when they are incubated with [ 35 S ]PAPS and normal sulfotransferases. These results suggest that in these patients the skeletal lesions are secondary to a defect in the synthesis of chondroitin sulfate involving specifically the sulfotransferase activity.

The Urinary Excretion of Heparan Sulfate by Juvenile- and Adult-Onset Diabetic Patients

The daily urinary excretions of total polymeric glycosaminoglycans and of polymeric heparan sulfate have been measured in the urine of juvenile-onset and adult-onset diabetics of both sexes and in those of normal controls. The results indicate that diabetic patients excrete more polymeric heparan sulfate than their controls, either in an absolute amount or as a percentage of the total glycosaminoglycans excreted. These results suggest that in the course of diabetes there is an increased degradation of heparan sulfate to large oligosaccharide fragments. These are excreted before being completely degraded to monosaccharides and inorganic sulfate.

Abnormally soluble collagen produced in fibroblasts cultures

Abnormally soluble collagen is synthesized in vitro not only by skin fibroblasts of Marfan patients but also by those of patients with Ehlers-Danlos type V and cutis laxa. The excessive solubility of collagen is corrected by the addition to the culture medium of a synthetic flavonoid, (+)-catechin.

Sulfated glycosaminoglycans synthesized by fibroblast, smooth muscle and endothelium-like cells grown in culture

Monolayer cultures of fibroblast, smooth muscle and endothelium-like cells incorporated 35SO2-(4) into glycosaminoglycans of the extracellular, pericellular and intracellular compartments. These glycosaminoglycans have been identified on the basis of electrophoretic mobility, enzymatic degradation with specific mucopolysaccharidases and by the type of degradation products formed. The sulfated glycosaminoglycans from the extracellular pool of the three cell types show a similar composition, while the intracellular and pericellular pools of the three cells have a different glycosaminoglycans composition. They differ in the relative proportion of heparitin sulfate and chondroitin sulfate and in the structure of isomeric chondroitin sulfate.

Antigenic determinants of two components of proteoglycan complex from bovine cartilage

The immunological properties of a glycoprotein fraction and of proteoglycan subunits obtained from bovine nasal cartilage by nondisruptive methods of isolation have been studied. Using the techniques of hemagglutination and hemagglutination inhibition, we found that the glycoprotein contains most of the species‐specific determinants, whereas the proteoglycan subunits contain most of the cross‐reacting ones.

The interaction of human plasma glycosaminoglycans with plasma lipoproteins. II. Hemagglutination studies.

Formalinized, tannic acid-treated sheep erythrocytes coated with low density lipoproteins (LSL) or apoprotein B (apo-B) are are agglutinated by anti-apo-B immunserum. Those coated with high density lipoproteins (HDL) or apoprotein A-I(apo-A-I) are agglutinated by anti-apo-A-I immunserum. These coated formocells have been used to study the interactions of lipoproteins and apoproteins with plasma glycosaminoglycans (GAG). The sulfate-rich species of plasma GAG agglutinates cells coated with LDL, HDL, apo-B, and apo-A-I at ionic concentrations above 0.15 M. The less-sulfated species of plasma GAG does not agglutinate the coated cells but inhibits the agglutination caused by the sulfate-rich species. Treatment of the sulfate-rich GAG with papain causes a reduction in molecular weight by one-half and also causes a loss of its agglutinating activity. These results suggest that the sulfate-rich plasma GAG, consisting of two glycan chains linked to a peptide backbone, cause agglutination by binding to two or more formocells. In contrast, the less-sulfated plasma GAG, consisting of single, short glycan chains, are incapable of causing agglutination but may prevent it by covering specific binding sites present on the coated cells.

Cultured fibroblasts of juvenile diabetics have excessively soluble pericellular collagen

Abstract The solubility of collagen synthesized by cultured skin fibroblasts derived from three patients affected by juvenile-onset diabetes mellitus and two normal controls has been compared. The diabetic collagen is much more soluble in 1.0 M NaCl pH 7.5 and in 0.5 M acetic acid. This excessive solubility is partially corrected by the addition to the diabetic cultures of a naturally-occuring flavonoid, (+)-catechin.

Cleavage of the (1→3)-2-acetamido-2-deoxy-β-d-glucopyranosyl linkage present in keratan sulfate. The A and B isoenzymes of human liver hexosaminidase (EC 3.2.1.30)

Abstract The disaccharide 2-acetamido-2-deoxy-β- d -glucopyranosyl-(1→3)- d -[1-3H]-galactitol, prepared from keratan sulfate, was rapidly hydrolyzed by the A and B isoenzymes of normal human liver hexosaminidase (EC 3.2.1.30), and by the B isoenzyme prepared from the liver of a patient who had died of Tay-Sachs disease. The disaccharide substrate was also hydrolyzed by extracts of normal, cultured-skin fibroblasts, and fibroblasts of patients with Tay-Sachs disease, whereas it was not hydrolyzed by fibroblast extracts of patients with Sandhoff disease. Thus, defective degradation of keratan sulfate, secondary to a defect of the β subunits present in the A and B isoenzymes of hexosaminidase, may contribute to the appearance of skeletal lesions in patients affected by Sandhoff disease.