Galactosamine Residues Research Articles

Pseudomonas aeruginosa bacteriophage phi PLS27-lipopolysaccharide interactions.

We investigated the phi PLS27 receptor in Pseudomonas aeruginosa strain PAO lipopolysaccharide (LPS) by analyzing a resistant mutant. This mutant, which was designated AK1282, had the most defective LPS yet reported for a P. aeruginosa rough mutant; this LPS contained only lipid A, 2-keto-3-deoxyoctonate, heptose, and alanine as major components. In addition, this LPS lacked galactosamine, which is present in the inner core of the LPS of other rough mutants. The loss of galactosamine but only a small decrease in the alanine content indicated that the core of strain PAO LPS differed from the core structure which has been suggested for the LPS of other well-characterized P. aeruginosa strains. Our analysis also indicated that galactosamine residues may be crucial for phi PLS27 receptor activity of the LPS. Electrodialysis of LPS and conversion to salt forms (sodium or triethylamine) influenced the phage-inactivating capacity of the LPS, as did the medium in which the inactivation occurred; experiments performed in 1/10-strength broth resulted in much lower PhI50 (concentration of LPS causing a 50% decrease in the titer of phage during 1 h of incubation at 37 degrees C) values than experiments performed in regular-strength broth. Sonication of the LPS also increased the phage-inactivating capacities of the LPS preparations.

Structure of Polygalactosamine Produced by <italic>Aspergillus parasiticus</italic>

The structure of polygalactosamine purified from the culture fluid of Aspergillus parasiticus AHU 7165 was studied. Partial acid hydrolysis of this polysaccharide, in which 55 to 65% of the monosaccharide residues were N-unsubstituted, gave a series of galactosamine oligosaccharides (dimer to hexamer) in a good yield. From the data on analyses of the polysaccharide and its oligosaccharides by gel filtration, periodate oxidation, methylation, and proton NMR measurement, the polysaccharide was characterized as a linear chain of alpha (1-4)-linked galactosamine residues. The N-unsubstituted galactosamine residues are probably distributed in a random fashion over the polysaccharide chain.

Age-related changes in the chemical composition of bovine articular cartilage. The structure of high-density proteoglycans.

Proteoglycans were extracted from the articular cartilage of foetal, calf and adult bovine metacarpal-phalangeal joints with 4m-guanidinium chloride. After extraction, the high-density proteoglycans (PG-I fractions) were prepared by sedimentation in two sequential CsCl-density-gradient procedures [Swann, Powell & Sotman (1979) J. Biol. Chem.254, 945-954]. The PG-I fractions from foetal, calf and adult tissues accounted for 75%, 52% and 46% respectively of the extracted components. The glucosamine, galactose, N-acetylneuraminic acid and protein contents increased with age. The overall amino acid compositions of PG-I fractions were similar. Fractionation of PG-I-fraction samples on a Bio-Gel A-50m column indicated that the molecular weight decreased with age. The PG-I fractions were specifically (3)H-labelled by treatment with galactose oxidase followed by reduction with NaB(3)H(4). The (3)H radioactivity was incorporated into both galactose and galactosamine residues of different carbohydrate side chains. The elution profiles of alkaline borohydride-treated foetal, calf and adult PG-I-fraction samples on a Sepharose 6B column showed that the molecular weights of chondroitin sulphate chains were 13500, 12000 and 10500 in foetal, calf and adult tissues respectively. Fractionation of the alkaline borohydride-treated foetal, calf and adult PG-I-fraction samples and (3)H-labelled calf and adult PG-I-fraction samples on a Bio-Gel P-10 column showed that there was an inverse relationship between the low-molecular-weight O-linked oligosaccharides and the higher-molecular-weight sialic acid-containing constituents at different ages. The oligosaccharide components of foetal, calf and adult PG-I-fraction samples represented 79%, 69% and 36% respectively of the total sialic acid content of the proteoglycans. Similarly in the (3)H-labelled calf and adult samples 75% and 30% of the total radioactivity were present in the oligosaccharide components respectively. Digestion with chondroitinase AC-II and infrared analyses showed that the PG-I-fraction F and C samples contained primarily chondroitin 4-sulphate chains whereas PG-I-fraction sample A was 6-sulphated. These studies show that the major proteoglycans (PG-I fractions) in the articular cartilage of foetal, calf and adult animals differ in the content, types and structure of the chondroitin sulphate, keratan sulphate and oligosaccharide constituents. These changes in proteoglycan structure reflect the gross age-related changes in the chemical composition of the tissue.

Cleavage of cell surface proteins by thrombin.

This study was based on our previous findings that the mitogenic action of thrombin on cultured fibroblasts can result from interaction of thrombin with the cell surface in the absence of internalization, and that the proteolytic activity of thrombin is required for stimulation of cell division. This prompted us to look for thrombin-mediated cleavages using 2-dimensional gel electrophoresis of labeled cell surface proteins. Surface membrane components were labeled by 3 procedures: 1) proteins were labeled by lactoperoxidase-catalyzed iodination using 125I-; 2) galactose and galactosamine residues of glycoproteins were oxidized with galactose oxidase and reduced with 3H-NaBH4; and 3) glycoproteins were metabolically labeled by incubating cells with 3H-fucose. labeling with the first 2 procedures was carried out after thrombin treatment; in contrast, cells metabolically labeled with 3H-fucose were subsequently treated with thrombin to look for proteolytic cleavages. Collectively, these studies indicated that only about 5 cell surface proteins were thrombin-sensitive, consistent with the high specificity of this protease. Each of the labeling procedures revealed a thrombin-sensitive cell surface glycoprotein which was identified as fibronectin by immunoprecipitation experiments. In addition, cell surface proteins of about 140K and 55K daltons were thrombin-sensitive. However, cell surface proteins of about 45K daltons and 130K to 150K daltons were increased after thrombin treatment. These experiments were conducted on an established line of Chinese hamster lung cells with the eventual goal of studying thrombin-mediated cleavages of cell surface proteins in a large number or in cloned populations derived from this line that are either responsive or unresponsive to the mitogenic action of thrombin. This approach should permit identification of proteolytic cleavages tha are necessary for thrombin-stimulated cell division.

Structure of the complex polysaccharide C-substance from Streptococcus pneumoniae type 1.

The complex cell-wall polysaccharide, C-substance, was isolated from Streptococcus pneumoniae type 1 and purified by DEAE-cellulose (HCO3(-) form) and Sephadex column chromatography. The complete structure of this antigen was obtained by the application of methylation and 1H NMR and 13C NMR spectroscopic techniques to a series of oligosaccharide fragments obtained by the selective degradation of the N-acetylated antigen. Native C-substance is composed of the following repeating unit: beta-D-Glup-1 leads to 3-alpha-AAT-Galp-1 leads to 4-alpha-D-GalNAcp-1 leads to 3-beta-D-GalNH2p-1 leads to 1'-ribitol-5-phosphate where AATGal is 2-acetamido-4-amino-2,4,6-trideoxygalactose. Phosphocholine substituents are situated at O(6) of the unacetylated galactosamine residues, and the repeating units are linked through a diphosphate ester from ribitol to O(6) of the beta-D-glucopyranose residue. This structure has also been shown to be common to C-substances prepared from a number of other pneumococcal types based on the criterion of their identical 13C NMR spectra.

Blood-group-ABH antigens of human erythrocytes. Quantitative studies on the distribution of H antigenic sites among different classes of membrane components.

The contribution of blood-group-active glycolipids and glycoproteins to the blood-group-ABH character of human erythrocytes was investigated. For that purpose the blood-group-H sites of human O cells were converted in vitro into group-A sites by transfer of alpha-N-acetyl-D-[14C]galactosamine residues with the aid of the blood-group-A gene-dependent alpha-N-acetylgalactosaminyl transferase prepared from human A1 plasma. Upon partition of the red cell membranes between water and organic solvent, about 5% of the label was found in the organic phase and about 20% in the water phase, thus reflecting the distribution of blood-group antigenic sites between glycosphingolipids with short carbohydrate chains and polyglycosylceramides, respectively. The fact that about 70% of the radioactivity remained tightly bound to the membranes and could only be released by treatment with pronase provided good evidence that the bulk of blood-group-H determinants is bound to glycoprotein material. Following these results it can thus be assumed that blood-group-ABH activity of human erythrocytes is determined preferentially by group-specific glycoproteins rather than glycolipids.

Attachment sites of carbohydrate moieties to peptide chain of bovine k-casein from normal milk.

The attachment site of the carbohydrate moiety to the peptide chain of normal κ-casein was investigated with κ-casein component P-6 containing the most carbohydrates. Three short glycopeptides 6-IB1, 6-IB2 − 1 and 6-IB2 − 2 were prepared by cyanogen bromide cleavage and digestion of proteases (pronase P and thermolysin). Glycopeptides 6-IB1, 6-IB2− 1 and 6-IB2 − 2 corresponded to residues 128–139 (Gly-Glu-Pro-Thr-Ser-Thr-Pro-Thr-Thr-Glu-Ala-Val), residues 128–132 (or 127–131) and residues 135–139 of κ-casein A, respectively, and contained threonine and/or serine, but not asparagine. Glycopeptide 6-IB1 was considered to have three carbohydrate chains because it contained three galactosamine residues. The results of alkali treatment of 6-IB, under reduction condition excluded serine residue as the binding site, and confirmed the existence of three binding sites in the carbohydrate moieties. The carbohydrate moiety was shown to attach to threonine residue No. 131 from analysis of 6-IB2− 1 and to threonine re...

Structural elucidation of the capsular polysaccharide antigen of Neisseria meningitidis serogroup Z using 13C nuclear magnetic resonance

The capsular polysaccharide antigen from Neisseriameningitidis serogroup Z contains equimolar quantities of 2-acetamido-2-deoxy-D-galactose, glycerol, and phosphate. Carbon-13 nuclear magnetic resonance indicates that the polysaccharide is composed by a repeating unit of 1′-O-2-acetamido-2-deoxy-α-D-galactopyranosyl-glycerol joined through phosphate diester groups at O-3′ of glycerol and O-3 of the galactosamine residue. Assignments of the signals in the carbon-13 nuclear magnetic resonance spectrum were made by consideration of the previously assigned signals of related monomers and oligomers. When applied to a Karplus type relationship the values of the 3J(13C–31P) coupling constants were consistent with the polysaccharide having an extended conformation.

Chondroitin SO4 catabolism in chick embryo chondrocytes

An enzyme preparation from cultured chick embryo vertebral chondrocytes attacks chondroitin SO4 oligosaccharides from the nonreducing terminal in a recycling pathway involving the sequential action of a beta-glucuronidase, a 4- or a 6-sulfatase, and a beta-N-acetylgalactosaminidase. The sequence is blocked by saccharo-1,4-lactone, an inhibitor of the beta-glucuronidase, or by 2-acetamido-2-deoxy-D-galactonolactone, an inhibitor of the beta-N-acetylgalactosaminidase. The level of 4-sulfatase activity is low relative to the other activities and limits the rate of catabolism of hybrid oligosaccharide structures containing both 6-sulfated galactosamine residues and 4-sulfated galactosamine residues. This results in the accumulation of shortened oligosaccharides, most of which have galactosamine-4-SO4 residues at their nonreducing terminals. In the presence of the lactone inhibitors, polymeric chondroitin SO4 is broken down by the enzyme preparation to oligosaccharides which are 10 to 15 monosaccharides long, indicating that degradation of chondroitin SO4 chains is initiated by an endoglycosidase which generates oligosaccharide substrates for the recycling exoglycosidase system.

Characterization of the surface glycoproteins of rat spermatozoa.

The present study investigates the macromolecular composition of a membrance fraction isolated from rat spermatozoa and uses specific biochemical probes to study the externally oriented plasma membrane glycoproteins of caput and cauda epididymal spermatozoa. A highly purified membrane fraction was isolated from rat cauda epididymal spermatozoa using sonication and differential centrifugation. SDS polyacrylamide gels of the isolated membrane fraction revealed a large number of Coomassie Blue staining bands (>25) and 5 PAS positive bands. The galactose oxidase-[3HI sodium borohydnide technique was employed on intact spermatozoa to radioactively label externally oriented plasma membrane glycoproteins possessing terminal galactose or galactosamine residues on their oligosaccharide chains. Cauda epididymal spermatozoa possess a 37,000 dalton glycoprotein on the cell surface which is labelled by this technique, but no such component is detected on caput epididymal spermatozoa. Similarly, radioactive labelling of surface sialo-glycoproteins by sequential treatment of spermatozoa with sodium metaperiodate and [3 HI sodium borohydride revealed that the 37,000 dalton glycoprotein could be labeled on cauda epididymal spermatozoa, but is not detected on caput epididymal spermatozoa. The significance of these results with respect to maturation of spermatozoa in the epididymis is discussed.

Properties of the human salivary aggregating factor for Leptotrichia buccalis cells

Leptotrichia buccalis cell-aggregating and blood-group substance appeared in the void-volume in gel filtration of human whole saliva on Sepharose 2B. The void-volume material remained in the sample gel in the disc gel electrophoresis at pH 9.5 and gave a single sedimentation peak in ultracentrifugation. Three preparations with different blood group substance activities, A, H and Le a, contained 18.2–22.2 per cent protein and 77.8–81.8 per cent carbohydrate. The carbohydrate was composed of glucosamine, galactosamine, fucose and galactose. Amino acid compositions of the three preparations were very similar, with the main acids being threonine, serine and proline followed by alanine and glycine. Total moles of the hydroxy-amino acids were about equal to the moles of galactosamine residue, except in the preparation from secretory saliva with blood-group A specificity. The findings suggest that the cell-aggregating factor is a high molecular-weight glycoprotein with blood-group activity. Isoelectric-focusing analysis revealed that the cell-aggregating factor was heterogeneous with respect to electrophoretic property. The cell-aggregating components were always accompanied by varying levels of blood-group activity.

The vitelline envelope of eggs from the giant keyhole limpet Megathura crenulata. I. Chemical composition and structural studies.

The egg vitelline envelope of the marine invertebrate Megathura crenulata is a glycoprotein composed of 37.3 mol % protein and 62.7 mol % carbohydrate. Of the total amino acid content, 61 mol % consists of a single amino acid, threonine. The carbohydrate content includes galactosamine, galactose, and fucose. The molar ratio of threonine to galactosamine is about 1:1. Most of the threonine residues are linked to galactosamine residues via O-glycosidic bonds. A single peptide that was purified following alkaline borohydride treatment of the vitelline envelope had the structure: Abu-Pro-Abu-(Abu6, Pro1, Thr1), where Abu is 2-aminobutyric acid. Several sugar residues have been isolated following the alkaline hydrolysis of the vitelline envelope that include an octasaccharide Gal4Fu4, an hexasaccharide Gal3Fu3, a trisaccharide Gal3, fucose, and galactose. It is proposed that the vitelline envelope of Megathura crenulata eggs is composed of polypeptide chains built to a large extent of closely spaced threonine residues. Almost every threonine residue is linked to a saccharide moiety.

Decomposition of blood group substance H in alkaline sodium borohydeide solution

The degradation of BGS H in alkaline NaBH4 solution is accompanied by the destruction of ∼40% of the galactosamine residues that are found in the node of the carbohydrate-peptide linkage. The other monosaccharides are not affected here, which makes it possible to use this procedure to obtain the carbohydrate chains of BGS with a minimum degree of degradation. Oligosaccharides with a variable molecular weight (from 500 to 3000 and higher) are formed as a result of the degradation, which indicates the great heterogeneity of the carbohydrate chains of the BGS. A number of the oligosaccharides was isolated in the pure state.

Antigenic polysaccharides of bacteria

The specific polysaccharide of Sh. boydii 6 is an acid hexosaminoglycan, the repeating unit of which is built from mannose, galactose, galactosamine, and glucuronic acid residues in a 2∶1∶1∶1 ratio.

Structure of the O-Glycosidically Linked Carbohydrate Units of Fetuin

Abstract Fetuin isolated from fetal calf serum was shown to contain 3 carbohydrate units which are attached to serine and threonine residues on the peptide chain and are distinct from the 3 asparagine-linked heteropolysaccharides previously reported to occur in this protein. The O-glycosidically linked saccharide units were obtained as reduced oligosaccharides after alkaline borohydride treatment of fetuin and as glycopeptides subsequent to pronase digestion of this protein. Gel filtration and Dowex 1 chromatography of the oligosaccharides resulted in the isolation of a tri- and a tetrasaccharide which were made up of sialic acid, galactose, and N-acetylgalactosaminitol residues and could be converted after selective removal of the sialic acid to the same disaccharide (galactosyl-N-acetylgalactosaminitol). Studies employing periodate oxidation and glycosidase digestion indicated that the structure of the tetrasaccharide was N-acetylneuraminyl-(2 →3)-β-d-galactopyranosyl-(1 →3)[N-acetylneuraminyl-(2 →6)]-N-acetylgalactosaminitol and that the trisaccharide differed only by the absence of the sialyl residue linked to the N-acetylgalactosaminitol. Glycopeptides containing the alkali-labile carbohydrate units were purified on Sephadex and diethylaminoethylcellulose columns. In this manner peptides distinguished by a high proline content and containing 1 tetrasaccharide, 1 trisaccharide, or 2 trisaccharides were obtained. Alkaline sulfite treatment of these glycopeptides indicated that 2 serine and 1 threonine residue in each fetuin molecule is involved in the attachment of carbohydrate units. The tetrasaccharide was found to be linked only to serine while attachment of trisaccharide units involved a serine and a threonine residue which were located in close proximity in the peptide chain. Smith periodate degradation of sialic acid-free glycopeptides resulted in removal of the galactose and yielded a product from which complete release of N-acetylgalactosamine was accomplished by digestion with α-N-acetylgalactosaminidase although no cleavage of this sugar was effected by the action of β-N-acetylhexosaminidase. This indicated that the alkali-labile units of fetuin are attached to the peptide chain by α-D-N-acetylgalactosaminyl-(1 →3)-serine (threonine) bonds. Of the total carbohydrate of fetuin 21%, including all three of the galactosamine residues, was shown to be located in the O-glycosidically linked units with the remainder occurring in the more alkali-stable N-glycosidically bound form. The alkaline sulfite treatment employed in this investigation produced in good yield the sulfonyl derivatives of the amino acids and amino sugar involved in the glycopeptide linkage. Since the cysteic acid and α-amino-β-hydroxybutyric acid obtained in this manner do not separate on the amino acid analyzer, a chromatographic procedure employing Dowex 1 was developed which permitted resolution of these two components.

The isolation and the structure of new glycopeptide from blood group substances

Three new glycopeptides with O-glycosidic and one glycopeptide with N-glycosylaminic carbohydrate-peptide linkages have been isolated after degradation of blood group substances (BGS). Their structure have been determined as O-(α-GalNAc)-Ser(I), O-(GalNAc)-(Pro)-Ser(II), O-(GalNAc 1 → 3 GalNAc)-(Thr-Ala)-Ser(III), N-(β-GlcNAc)-Asn(IV). The isolation of glycopeptide I confirmed α-configuration of O-glycosidic carbohydrate-peptide bonds. The structure of glycopeptide III with two galactosamine residues is in accordance with the data on the presence of hexosamine core of BGS carbohydrate chains.

Physico-chemical characteristics and regional distribution studies of GP-350, a soluble sialoglycoprotein from brain.

AbstractThe apparent molecular weight of GP‐350, a sialoglycoprotein from calf and rat brain, has been determined by SDS‐polyacrylamide gel electrophoresis. The electrophoretic mobility corresponds to the mobility of a polypeptide with a molecular weight of 11,600 ± 200. On this basis it can be calculated that only one sialic acid residue is present/GP‐350 molecule.From isoelectric focusing experiments it appeared that the isoelectric point of GP‐350 is about 2. The determination of the amide content of the polypeptide chain showed that out of 22.0 acidic amino acid residues of glutamic acid and aspartic acid, only 4.9 residues are amidated. The total amount of the basic amino acid residues lysine and arginine, is 6.5. So, per molecule GP‐350 10.6 acidic amino acid residues are not counteracted by basic amino acid residues. The surplus of the acidic amino acid residues as well as sialic acid result in the pronounced acidic character of GP‐350. This fact is supported by the electrophoretic experiments.The carbohydrate‐polypeptide linkage type has been studied by alkaline sodium borohydride treatment. Two thirds of all the galactosamine was destroyed, whereas the amount of glucosamine remained the same. Amino acid analysis indicated a decrease in serine and threonine with a concomitant small increase in alanine. These data point to the occurrence of linkages between the carbohydrate chain and the polypeptide core of the galactosamineserine or –threonine type. Per molecule GP‐350 about two residues of galactosamine are destroyed, indicating that two carbohydrate chains of this binding type are present. Only one of these chains can be terminated by a sialic acid residue. The other carbohydrate chain may be terminated by fucose.Regional distribution studies showed the presence of GP‐350 in all brain areas studied; in relatively large amounts in the regions rich in ganglia such as caudate nucleus, cerebellar grey matter, pons and medulla oblongata, and in relatively small amounts in the regions poor in ganglia such as corpus callosum, cerebral white, cerebral grey and cerebellar white matter.GP‐350 is also present in the pituitary gland. In the cerebrospinal fluid a glycoprotein is present with the same electrophoretic mobility as GP‐350. However, this glycoprotein gave no precipitin reaction with GP‐350 specific antiserum. Moreover, the amino acid composition was quite different from that of GP‐350.Subcellular distribution study revealed that GP‐350 is present in the soluble cell fraction and in the synaptosomal membrane fraction, whereas it is absent from the purified nuclei, mitochondria, myelin, and also from the microsomal fraction.

Studies on the Carbohydrate Units of Thyroglobulin: EVALUATION OF THEIR MICROHETEROGENEITY IN THE HUMAN AND CALF PROTEINS

Information has been obtained in regard to the carbohydrate units of human and calf thyroglobulin through a study of the glycopeptides obtained after extensive Pronase digestion of these proteins. Human thyroglobulin was shown to contain, as has been previously reported for the calf protein, carbohydrate units consisting only of mannose and N-acetylglucosamine (unit A) as well as more complex units consisting of mannose, Nacetylglucosamine, galactose, N-acetylneuraminic acid, and fucose (unit B). The units A of the human and calf thyroglobulin were similar, consisting on an average of 8 to 9 residues of mannose and 2 of N-acetylglucosamine. There were 7 to 8 units of this type in the human protein and 5 to 6 in the calf. Chromatography of human and calf unit A glycopeptides on Dowex 50-X2 revealed extensive microheterogeneity, with the mannose content varying from 5 to 11 residues per unit, but with the glucosamine content remaining constant at 2 residues in each. The average unit B of the human thyroglobulin was found to consist of 3 mannose, 4 N-acetylglucosamine, 2 to 3 galactose, 1 fucose, and 1 sialic acid residues. It differed from that previously reported for the calf in containing fewer residues of sialic acid, galactose, and glucosamine. There were found to be 22 units of the B type in the human thyroglobulin compared with 13 to 14 of these units in the calf protein. The human unit B was also found to exhibit pronounced microheterogeneity, as was revealed by fractionation of its glycopeptides on DEAE-cellulose. The variation was primarily evident in the sialic acid, galactose, and glucosamine residues, with the mannose of the core portion remaining constant at 3 residues. The galactosamine residues of the human thyroglobulin were shown, by density gradient centrifugation, to be an integral part of this protein and to occur in units distinct from carbohydrate units A and B, which have been designated as unit C. Alkaline sulfite treatment of glycopeptides containing galactosamine suggested that the C type of unit is linked to serine and threonine residues by O-glycosidic bonds, in contrast to the glycosylamine type of linkage to asparagine which attaches units A and B to the peptide chain. During Pronase digestion, some of the glycopeptide bonds linking unit A were split, yielding neutral oligosaccharides containing 7 to 8 mannose and 2 N-acetylglucosamine residues, with 1 of the latter in a reducing terminal position.

The composition of beta-lactamase I and beta-lactamase II from Bacillus cereus 569-H.

1. Crystalline beta-lactamase I from Bacillus cereus 569/H yielded only amino acids on acid hydrolysis, but crystalline beta-lactamase II from the same organism yielded also substantial quantities of neutral sugars and amino sugars. 2. Analysis with an amino acid analyser indicated that the two enzymes were similar though not identical in overall amino acid composition. Analysis of neutral and amino sugars as their silyl derivatives by gas-liquid chromatography showed that the carbohydrate moiety of beta-lactamase II contained residues of glucose, galactose, mannose, fucose, glucosamine and galactosamine. 3. After oxidation and hydrolysis both beta-lactamases gave small amounts of cysteic acid. After treatment of inactive Zn(2+)-free beta-lactamase II with N-ethylmaleimide or iodoacetate enzymic activity was not restored by the addition of Zn(2+).

Structure of Dermatan Sulfate: III. THE HYBRID STRUCTURE OF DERMATAN SULFATE FROM UMBILICAL CORD

Abstract The properties of dermatan sulfate, isolated from umbilical cord, have been compared with those of pig skin dermatan sulfate. Like the latter polysaccharide, umbilical cord dermatan sulfate was found to be a hybrid containing both l-iduronic and d-glucuronic acid; whereas the galactosamine units of pig skin dermatan sulfate were exclusively 4-sulfated, the umbilical cord polysaccharide contained a considerable proportion of 6-sulfated galactosamine residues. The latter components were preferentially located adjacent to d-glucuronic acid moieties, while 4-sulfated galactosamine units were prevalent in l-iduronic acid-containing sections of the molecule. Evidence was obtained, however, indicating that some of the galactosamine residues adjacent to l-iduronic acid moieties were 6-sulfated. This conclusion was based on the characterization of hybrid octasaccharides obtained after degradation with testicular hyaluronidase. These octasaccharides which all had the monosaccharide sequence shown below contained 2 chondroitin 6-sulfate disaccharide units located in the reducing and nonreducing termini, and 2 l-iduronic acid-containing internal disaccharide units which were largely 4-sulfated. Approximately one-third of the latter disaccharides, however, appeared to be sulfated in position 6. [see PDF for sequence]