Alpha-galactose Residues Research Articles

Physicochemical properties of pectins from okra ( Abelmoschus esculentus (L.) Moench)

Okra pectin obtained by hot buffer extraction (HBSS) consists of an unusual pectic rhamnogalacturonan I structure in which acetyl groups and alpha galactose residues are substituted on rhamnose residues within the backbone. The okra Chelating agent Soluble Solids (CHSS) pectin consists of slightly different structures since relatively more homogalacturonan is present within the macromolecule and the rhamnogalacturonan I segments carry slightly longer side chains. The rheological properties of both okra pectins were examined under various conditions in order to understand the unusual slimy behaviour of okra pectins. The viscosity of the okra HBSS pectin was 5–8 times higher than the viscosity of the okra CHSS pectin. The okra HBSS pectin showed an elastic behaviour ( G′ > G″) over a wide range of frequencies (10 −1–10 Hz), at a strain of 10%, while okra CHSS and saponified okra HBSS/CHSS pectin showed predominantly viscous responses ( G′ < G″) over the same frequency range. The results suggest that the structural variation within the okra pectins greatly affect their rheological behaviour and it is suggested that acetylation of the pectin plays an important role through hydrophobic associations. Dynamic light scattering was used to study the association behaviour of both okra pectins at low concentration (0.001–0.1% w/w). Results showed that the saponified okra pectins did not exhibit a tendency to aggregate in the concentration range studied, whereas both non saponified samples showed a substantial degree of association. These results suggest that the unusual slimy behaviour of the non saponified samples may be related to the tendency of these pectins to associate, driven by hydrophobic interactions.

Identification of high-mannose and multiantennary complex-type N-linked glycans containing α-galactose epitopes from Nurse shark IgM heavy chain

MALDI-TOF mass spectrometry, negative ion nano-electrospray MS/MS and exoglycosidase digestion were used to identify 36 N-linked glycans from 19S IgM heavy chain derived from the nurse shark (Ginglymostoma cirratum). The major glycan was the high-mannose compound, Man(6)GlcNAc(2) accompanied by small amounts of Man(5)GlcNAc(2), Man(7)GlcNAc(2) and Man(8)GlcNAc(2). Bi- and tri-antennary (isomer with a branched 3-antenna) complex-type glycans were also abundant, most contained a bisecting GlcNAc residue (beta1-->4-linked to the central mannose) and with varying numbers of alpha-galactose residues capping the antennae. Small amounts of monosialylated glycans were also found. This appears to be the first comprehensive study of glycosylation in this species of animal. The glycosylation pattern has implications for the mechanism of activation of the complement system by nurse shark IgM.

成熟・受精・発生過程におけるブタ卵細胞膜のα-ガラクトース残基の発現について

成熟・受精・発生過程におけるブタ卵細胞膜のα-ガラクトース残基の発現について

Lectin/glycan interactions play a role in recognition in a coral/dinoflagellate symbiosis

Recognition is an important stage in the establishment of highly specific mutualistic associations. Yet, for the majority of symbioses, very few of the mechanisms involved in recognition and specificity are known. In this study, we provide evidence for a recognition mechanism at the onset of symbiosis between larvae of the coral Fungia scutaria and their endosymbiotic dinoflagellate algae. This recognition step occurs during initial cellular contact between the symbiotic partners through a lectin/glycan interaction. We determined that an intact algal cell surface was required for successful infection of F. scutaria larvae. Modification of the algal cell surface by enzymatic digestion with trypsin or N-glycosidase significantly reduced infection success, and implicated algal cell surface glycans in recognition. Using flow cytometry, alpha-mannose/alpha-glucose and alpha-galactose residues were identified as potential recognition ligands on the algal cell surface. Finally, inhibition of these cell surface glycans significantly reduced infection of F. scutaria larvae by the algae. These data provide evidence that the algal cell surface contains glycan ligands, such as alpha-mannose/alpha-glucose and alpha-galactose, which play a role in recognition during initial contact at the onset of symbiosis with F. scutaria larvae.

The identification and characterization of xenoantigenic nonhuman carbohydrate sequences in membrane proteins from porcine kidney

The immunogenic nonhuman carbohydrate sequences in membrane proteins from porcine kidney were identified and characterized using MALDI-TOF MS and ESI-QTOF-MS. The MALDI profile, investigated by incubation with exoglycosidases, showed a series of about 40 carbohydrates that were identified as high mannose glycans (Man(3-9)GlcNAc2) and complex bi-, tri-, and tetra-antennary glycans with and without core fucose. The antennae of many of the complex glycans were terminated with alpha-galactose residues, with the numbers of these residues ranging from one up to the number of antennae. Negative ion ESI-MS/MS spectra confirmed the location of the alpha-galactose residues on the ends of the antennae. This total glycan profile of the membrane proteins from porcine kidney will thus provide important information for the study of molecular interactions between antigenic carbohydrates and proteins in xenotransplantation.

Ultrastructural localization of α-galactose-containing glycoconjugates in the rat vomeronasal organ

Binding sites of Griffonia simplicifolia I-B4 isolectin (GS-I-B4), which recognizes terminal alpha-galactose residues of glycoconjugates, were examined in the juxtaluminal region of the rat vomeronasal sensory epithelium and its associated glands of the vomeronasal organ, using a lectin cytochemical technique. Lowicryl K4M-embedded ultra-thin sections, which were treated successively with biotinylated GS-I-B4 and streptavidin-conjugated 10 nm colloidal gold particles, were observed under a transmission electron microscope. Colloidal gold particles, which reflect the presence of terminal alpha-galactose-containing glycoconjugates, were present in vomeronasal receptor neurons in the sensory epithelium and secretory granules of acinar cells of associated glands of the epithelium. Quantitative analysis demonstrated that the density of colloidal gold particles associated with sensory cell microvilli that projected from dendritic endings of vomeronasal neurons was considerably higher than that of microvilli that projected from neighboring sustentacular cells. The same was true for the apical cytoplasms of these cells just below the microvilli. These results suggest that of the sensory microvilli and dendritic endings contained a much larger amount of the alpha-galactose-containing glycoconjugates, compared with those in sustentacular microvilli. Further, biochemical analyses demonstrated several vomeronasal organ-specific glycoproteins with terminal alpha-galactose.

Crystallization and preliminary X-ray crystallographic studies of rice alpha-galactosidase.

alpha-Galactosidases catalyze the hydrolysis of galactooligosaccharides and galactopolysaccharides to alpha-galactose residues and are widely distributed in microorganisms, plants and animals. alpha-Galactosidase from rice (Oryza sativa L. ssp. japonica) was crystallized by the hanging-drop vapour-diffusion method. The crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 63.1, b = 71.3, c = 85.6 A, and diffract beyond 1.9 A resolution.

Structural study on the carbohydrate moiety of calf intestinal alkaline phosphatase.

Surprisingly alkaline phosphatase (AP) (EC 3.1.3.1) of calf intestine is found in large amounts, e.g. 80%, within chyme. Most of the enzyme is present as a mixture of four differently hydrophobic anchor-bearing forms and only the minor part is present as an anchorless enzyme. To investigate whether changes in the N-glycosylation pattern are signals responsible for large-scale liberation from mucosa into chyme, the glycans of the two potential glycosylation sites predicted from cDNA were investigated by matrix-assisted laser desorption/ionization and electrospray ionization mass spectrometry in combination with exoglycosidase treatment after tryptic digestion and reversed-phase chromatography. The glycans linked to Asn249 are at least eight different, mainly non-fucosylated, biantennary or triantennary structures with a bisecting N-acetylglucosamine. For the most abundant glycopeptide (40%) the following glycan structure is proposed: [carbostructure: see text]. The glycans linked to Asn410 are a mixture of at least nine, mainly tetraantennary, fucosylated structures with a bisecting N-acetylglucosamine. For the most abundant glycopeptide (35%) the following glycan structure is proposed: [carbostructure: see text]. For the structures the linkage data were deduced from the reported specificities of the exoglycosidases used and the specificities of the transglycosidases active in biosynthesis. The majority of glycans are capped by alpha-galactose residues at their non-reducing termini. In contrast to the glycans linked to other AP isoenzymes, no sialylation was observed. Glycopeptide 'mass fingerprints' of both glycosylation sites and glycan contents do not differ between AP from mucosa and chyme. These results suggest that the observed large-scale liberation of vesicle-bound glycosylphosphatidylinositol (GPI)-anchored AP from mucosa into chyme is unlikely to be mediated by alteration of glycan structures of the AP investigated. Rather, the exocytotic vesicle formation seems to be mediated by the controlled organization of the raft structures embedding GPI-AP. (c) 2001 John Wiley & Sons, Ltd.

Structure and Anticoagulant Activity of Sulfated Galactans: ISOLATION OF A UNIQUE SULFATED GALACTAN FROM THE RED ALGAEBOTRYOCLADIA OCCIDENTALIS AND COMPARISON OF ITS ANTICOAGULANT ACTION WITH THAT OF SULFATED GALACTANS FROM INVERTEBRATES

We have characterized the structure of a sulfated d-galactan from the red algae Botryocladia occidentalis. The following repeating structure (-4-alpha-d-Galp-1-->3-beta-d-Galp-1-->) was found for this polysaccharide, but with a variable sulfation pattern. Clearly one-third of the total alpha-units are 2,3-di-O-sulfated and another one-third are 2-O-sulfated. The algal sulfated d-galactan has a potent anticoagulant activity (similar potency as unfractionated heparin) due to enhanced inhibition of thrombin and factor Xa by antithrombin and/or heparin cofactor II. We also extended the experiments to several sulfated polysaccharides from marine invertebrates with simple structures, composed of a single repeating structure. A 2-O- or 3-O-sulfated l-galactan (as well as a 2-O-sulfated l-fucan) has a weak anticoagulant action when compared with the potent action of the algal sulfated d-galactan. Possibly, the addition of two sulfate esters to a single alpha-galactose residue has an "amplifying effect" on the anticoagulant action, which cannot be totally ascribed to the increased charge density of the polymer. These results indicate that the wide diversity of polysaccharides from marine alga and invertebrates is a useful tool to elucidate structure/anticoagulant activity relationships.

Differential expression of glycoside residues in the mammalian zona pellucida.

The mammalian zona pellucida is an extracellular matrix surrounding the oocyte, and is composed of three major glycoproteins, ZP1, ZP2, and ZP3. Previous studies have suggested that the sperm receptor activity of the zona pellucida resides in specific oligosaccharide chains on the ZP3 glycoprotein. However, the nature of the terminal monosaccharide(s) on these glycosidic chains to which sperm bind is a matter of active debate. Evidence has been presented to support a role for at least three distinct monosaccharides in sperm binding, alpha-galactose, L-fucose on Lewis X structures, and beta-N-acetylglucosamine. Previous studies have shown that beta-N-acetylglucosamine is uniformly distributed throughout the zona matrix. In this study, we have investigated the expression and distribution of alpha-galactose and fucose moieties during the maturation of the zona pellucida in mouse, rat, and hamster. Interestingly, alpha-galactose residues are expressed only during later stages of zona secretion and, consequently, are confined to the inner portions of the mature zona pellucida in mouse and rat. In hamster, alpha-galactose residues are only detectable in the zona pellucida of ovulated eggs, and are not found in ovarian oocytes. Fucosyl residues linked to Lewis X glycosides are not detectable at any stage of zona maturation in these three species, whereas fucose linked to N-linked core oligosaccharides are present throughout the zona. These studies indicate a previously unappreciated heterogeneity in the composition of zona glycosides. The specific localization of alpha-galactose residues to the inner portions of the zona matrix suggest a role in the later stages of sperm penetration through the zona. Finally, due to their absence from the zona surface, alpha-galactose and Lewis X fucosyl residues are not likely to be mediators of primary sperm binding.

Functional Evidence for UDP-galactose Transporter inSaccharomyces cerevisiae through the in Vivo Galactosylation and in Vitro Transport Assay

The oligosaccharide profiles in glycoproteins are determined by a series of processing reactions catalyzed by Golgi glycosyltransferases and glycosidases. Recently in vivo galactose incorporation in Saccharomyces cerevisiae has been demonstrated through the expression of human beta-1,4-galactosyltransferase in an alg1 mutant, suggesting the presence of a UDP-galactose transporter in S. cerevisiae (Schwientek, T., Narimatsu, H., and Ernst, J. F. (1996) J. Biol. Chem. 271, 3398-3405). However, this is quite unexpected, because S. cerevisiae does not have galactose residues in its glycoproteins. To address this question we have constructed S. cerevisiae mnn1 mutant strains expressing Schizosaccharomyces pombe alpha-1,2-galactosyltransferase. The mnn1 mutant of S. cerevisiae provides endogenous acceptors for galactose transfer by the expressed alpha-1,2-galactosyltransferase. We present here three lines of evidences for the existence of UDP-galactose transporter in S. cerevisiae. (i) About 15-20% of the total transformed mnn1 cells grown in a galactose medium were stained with fluorescein isothiocyanate-conjugated alpha-galactose-specific lectin, indicating the presence of alpha-galactose residues on the cell surface. (ii) Galactomannan proteins can be precipitated with agarose-immobilized alpha-galactose-specific lectin from a whole cell lysate prepared from transformed mnn1 cells grown in a galactose medium. (iii) The presence of UDP-galactose transporter was demonstrated by direct transport assay. This transport in S. cerevisiae is dependent on time, temperature, and protein concentration and is inhibited by nucleotide monophosphate and Triton X-100. The overall UDP-galactose transport in S. cerevisiae is comparable with that in S. pombe, indicating a more or less similar reaction velocity, while the rate of GDP-mannose transport is higher in S. pombe than in S. cerevisiae.

The Glycosylation of the Influenza A Virus Hemagglutinin by Mammalian Cells: A SITE-SPECIFIC STUDY

We have characterized the glycans at individual sites on the hemagglutinin of three influenza A variants to obtain information on the role of cell-specific glycosylation in determining the receptor binding properties of this virus. The variants differ in whether they have a glycosylation site at residue 129 on the tip of the hemagglutinin and whether amino acid 184 (near to the receptor binding site) is His or Asn. We found that all sites on each variant are glycosylated in Madin-Darby bovine kidney cells, that the glycosylation is site-specific, and that the glycans at the same site in each variant are highly similar. One site that is buried in the hemagglutinin trimer contains only oligomannose glycans. The remaining sites carry complex glycans of increasing size as the distance of the site from the viral membrane decreases. Most of these complex glycans are terminated with alpha-galactose residues, a consequence in bovine cells of the removal of terminal sialic acids by the viral neuraminidase. Although the glycans at residue 129 are among the smallest on the molecule, they are large enough to reach the receptor binding pocket on their own and adjacent monomers. The results suggest that the reduction in receptor binding observed with Madin-Darby bovine kidney cell-grown virus is due to the combined effect of large complex glycans at the tip of the hemagglutinin and a His to Asn substitution close to the receptor binding pocket.

Resolution of sensory and mucoid glycoconjugates with terminal alpha-galactose residues in the mucomicrovillar complex of the vomeronasal sensory epithelium by dual confocal laser scanning microscopy.

The organization of the mucomicrovillar complex of the vomeronasal sensory epithelium of adult rats was examined using confocal laser scanning microscopy. In specimens labeled with the FITC-conjugated isolectin B4 of Bandeiraea simplicifolia, which recognizes terminal alpha-galactose sugar residues of glycoconjugates, we demonstrated that the mucomicrovillar complex was composed of islet-like structures with a high-density alpha-galactose core. The mucomicrovillar complex was further resolved into sensory and mucoid components in double-labeling and dual scanning experiments. The sensory component, which consists of the dendritic terminals of olfactory marker protein-immunoreactive vomeronasal receptor neurons, contained cytosolic glycoconjugates with terminal alpha-galactose sugar residues. The extracellular mucoid component consisted of glycoconjugates containing terminal alpha-galactose derived from the glands associated with the vomeronasal organ. These results demonstrated the complex microchemical organization of the sensory and mucoid components of the mucomicrovillar complex.

Site-specific glycosylation of recombinant rat and human soluble CD4 variants expressed in Chinese hamster ovary cells.

The rat and human forms of the T-cell surface glycoprotein CD4 share a common glycosylation site at the Asn270/271 position but differ with respect to the locations of the second glycosylation sites at Asn159 (rat) and Asn300 (human). The glycosylation of soluble recombinant forms of human and rat CD4 (sCD4) expressed in Chinese hamster ovary cells has been characterized. The most obvious differences between the rat and human sCD4 oligosaccharides were the greater abundance of oligomannose and hybrid oligosaccharides on rat sCD4 and the presence of oligosaccharides carrying a terminal alpha-galactose residue on human sCD4. This is the first report of the occurrence of alpha-galactose residues on a glycoprotein expressed in Chinese hamster ovary cells. Comparison of mutant rat sCD4 molecules with single glycosylation sites and glycopeptides indicated that site-specific and independent processing occurred at each glycosylation site. The glycosylation at the conserved site at Asn270 of rat sCD4 was identical to that seen for the equivalent site in human sCD4, and the oligomannose and hybrid structures were restricted to the nonconserved site at Asn159 in rat sCD4. However, there was more oligosaccharide processing at this site in a truncated form of rat sCD4 consisting of the two NH2-terminal domains. These results indicate that not only the local three-dimensional structure but also domain interactions can influence the processing at individual glycosylation sites.

Detailed structural analysis of asparagine-linked oligosaccharides of the nicotinic acetylcholine receptor from Torpedo californica.

The structures of the major oligosaccharide moieties of the nicotinic acetylcholine receptor (AcChoR) protein from Torpedo californica have been reported [Nomoto, H., Takahashi, N., Nagaki, Y., Endo, S., Arata, Y. and Hayashi, K. (1986) Eur. J. Biochem. 157, 233-242] to be high-mannose types. Here we report detailed analyses of the structures of the remaining oligosaccharides in this receptor. The sialylated oligosaccharides released by glycopeptidase (almond) digestion were separated according to the number of sialic acid residues using high-performance anion-exchange chromatography with pulsed amperometric detection. After removal of sialic acid from each fraction, the resulting neutral oligosaccharides were separately pyridylaminated and were analyzed by a combination of sequential exoglycosidase digestion and HPLC, then identified on a two-dimensional sugar map. The structures of two desialylated pyridylamino-oligosaccharides were further analyzed by high-resolution proton NMR. Each oligosaccharide was composed of species containing varying numbers of sialic acids. The desialylated complex-type oligosaccharides of AcChoR consisted of ten, eight and one different biantennary, triantennary and tetraantennary oligosaccharide, respectively. The biantennary oligosaccharides were divided into two groups; oligosaccharides with fucose at the proximal N-acetylglucosamine (six varieties) and oligosaccharides without fucose (four varieties). Each group consisted of species differing in the number of terminal galactose residues. The major component of the biantennary oligosaccharides had two galactose residues at the non-reducing termini. The terminal alpha-galactose residue(s) linked to C3 of beta-galactose were found in the fucose-containing biantennary oligosaccharides (two varieties). The triantennary oligosaccharides were also divided into two groups; oligosaccharides with (four varieties) and without (four varieties) besecting N-acetylglucosamine. These groups were composed of species differing in the number of terminal galactose residues. The major component of the triantennary oligosaccharides was fully galactosylated with three galactose residues. An unusual group, Gal beta 1-3GlcNAc, was present in low levels in the triantennary oligosaccharides. In contrast, the tetraantennary oligosaccharide was composed of only one species, which is fully galactosylated with four galactose residues.

Comparison of theMaclura pomifera lectin-binding glycoprotein in late fetal and adult rat lung

The lectin, Maclura pomifera agglutinin (MPA), binds to alpha-galactose residues of glycoproteins on the apical surface of type II alveolar cells. It has recently been shown to bind to macrophages. We isolated the cell surface glycoprotein, which binds the MPA lectin, from fetal and adult rat whole lung to determine if changes in this glycoprotein occur during development from fetal to adult life. The glycoprotein was purified from whole lung cell membranes by lectin affinity chromatography that resulted in 10(5)-fold enrichment. The MPA binding glycoproteins from both fetal and adult lung had the same apparent molecular weight of 170 kD as determined by sodium dodecylsulfate-polyacrylamide gel electrophoresis. Amino acid analysis revealed similar composition of the fetal and adult proteins. Two-dimensional peptide maps of the 170 kD proteins isolated from fetal and adult lung were also similar. These data indicate that the glycoprotein that binds MPA to lung cell membranes does not change during this stage of development. Our method for the isolation of this glycoprotein can be used for the generation of antibodies or other molecular probes for further study of this protein.

Presence of an α-galactolipid on the cell surfaces of endothelial cells of human kidney

The presence of an alpha-galactolipid was investigated with a peroxidase-labelled lectin from Griffonia simplicifolia (GSA-I) with specific binding for terminal alpha-D-galactose residues. Normal kidney tissue was obtained from patients undergoing nephrectomy for renal neoplasms. For light microscopy, tissue was snap-frozen; 4 microns-thick sections were briefly fixed in paraformaldehyde and incubated with GSA (0.025 mg ml-1). The peroxidase activity was developed with 3-amino-9-ethylcarbazole. Adjacent sections were stained at the same time after lipid extraction with 3:1 (v/v) chloroform/methanol. For electron microscopy, 0.2-0.5 mm-thick paraformaldehyde-fixed blocks, with or without lipid extraction, were stained with peroxidase-labelled GSA. The label was developed with diaminobenzidine and osmium tetroxide. Some structures, such as tubular epithelia, stained both in lipid-extracted and non-extracted tissues, suggesting that glycoproteins were most likely involved. In addition, tissue stained immediately after fixation showed GSA reactivity on endothelial cell surfaces of intertubular capillaries and larger vessels. In lipid-extracted tissues, however, tubular epithelium was still positive for GSA but endothelial cells failed to stain. These findings suggest that a glycolipid, bearing a terminal alpha-galactose residue, is present on the endothelial cells in human kidney and possibly on tubular epithelia. Our data may explain the preferential storage of alpha-galactolipid in endothelial cells of patients with Fabry's disease and other biological phenomena such as Escherichia coli adhesion.

Carbohydrate moieties of the basal lamina: their role in attachment and spreading of basal corneal epithelial cells.

Three lectins, Wheat germ agglutinin, succinyl Concanavalin A and Ricinus communis agglutinin were used to block specific sugar moieties in the basal lamina. Corneal epithelial basal cells were plated onto freshly denuded basal lamina. Attachment was studied by quantifying the adherence of prelabeled cells and by examining attachment sites using transmission electron microscopy. Spreading was examined using scanning electron microscopy. Attachment of the cells occurred within 15 min and spreading was apparent after 45 min. Both Wheat germ agglutinin and beta-N-acetylglucosaminidase inhibited cellular attachment. Succinyl Concanavalin A and Ricinus Communis agglutinin permitted attachment, but inhibited extensive cellular spreading. The results indicate that the attachment of basal cells is dependent on N-acetylglucosamine residues, and spreading is mediated by alpha methylmannoside, glucose, and galactose residues.

Glycoconjugate with terminal alpha galactose. A property common to basal cells and a subpopulation of columnar cells of numerous epithelia in mouse and rat.

Glycoconjugates associated with the basal cell layer of various types of epithelia in the mouse and rat were examined histochemically with a battery of lectin-horseradish peroxidase (HRP) conjugates of differing sugar binding specificities. Basal cells in paraffin sections of composite tissue blocks stained with an isolectin from Griffonia simplicifolia (GSA I-B4) specific for terminal alpha-galactose residues but failed to react with the other lectins. Basal cells in epithelium lining striated and excretory ducts of salivary and lacrimal glands, tongue, esophagus, trachea, renal calyx, ureter, urinary bladder, urethra, epididymis and vas deferens stained selectively and intensely for content of a glycoconjugate with terminal alpha-galactose. This galacto-conjugate appeared associated with the plasmalemma of basal cells. Basal cells with a galactocalyx formed an intermittent to continuous layer generally increasing in prevalence distally in glandular duct systems. A minor population of pyramido-columnar cells with cytosolic GSA I-B4 reactivity occurred in striated ducts and appeared less numerous in intralobular excretory ducts and more prevalent in extraglandular ducts. In trachea and renal pelvis, the GSA I-B4 positive cell profiles ranged from low cuboidal to tall pyramidal in contour, but the latter appeared not to reach the lumen. In contrast, no GSA I-B4 positive basal cells were seen in any segment of the pancreatic or bile ducts or in the epithelium of the gastrointestinal tract. These findings suggest that the basal cells found in similar sites in different epithelia and possessing in common a unique alpha-galactoconjugate may function in a manner common to all and not simply in providing progenitor cells for epithelial renewal.(ABSTRACT TRUNCATED AT 250 WORDS)

Activation of mouse peritoneal macrophages alters the structure and surface expression of protein-bound lactosaminoglycans.

We have begun to analyze and compare the surface carbohydrates present on populations of resident and activated mouse peritoneal macrophages. The activated macrophage populations studied include TG-elicited macrophages, BCG-activated macrophages, and resident macrophages cultured for 24 hr in the presence of lymphokines or heterologous serum. Analysis of glycopeptides generated by pronase digestion of surface glycoproteins labeled by the neuraminidase/galactose oxidase/NaB3H4 method indicates that the macrophage surface contains a class of high m.w. carbohydrates susceptible to degradation by endo-beta-galactosidase, lactosaminoglycans. These lactosaminoglycans are sialylated type 2 carbohydrates containing the repeating lactosamine disaccharide Gal beta 1-4GlcNAc as well as fucose residues. Macrophage activation was observed to markedly alter surface lactosaminoglycans. The alterations observed include 1) an increase in surface expression as determined by both an increase in neuraminidase/galactose oxidase/NaB3H4 labeling and by the ability of activated but not resident macrophages to bind I antibodies as assayed by indirect immunofluorescent surface staining, 2) the addition of alpha-galactose residues, and 3) an increase in GlcNAc beta 1-6Gal branching as indicated by an increased resistance to endo-beta-galactosidase degradation and by the ability of activated macrophages to bind I antibodies. These observations demonstrate that macrophage activation results in specific and substantial alterations in protein-bound surface carbohydrates.