Effects of Trypsin and Plasmin Treatment of Myelin on the Myelin-Associated Glycoprotein and Basic Protein

Human and rat myelin preparations were incubated with varying concentrations of trypsin and plasmin to determine the effects of these proteolytic enzymes on myelin-associated glycoprotein (MAG), basic protein, and other myelin proteins and to compare the effects with those of the neutral protease that was reported to be endogenous in myelin. Basic protein was most susceptible to degradation by both trypsin and plasmin, whereas MAG was relatively resistant to their actions. Under the assay conditions used, the highest concentrations of trypsin and plasmin degraded greater than 80% of the basic protein but less than 30% of the MAG, and lower concentrations caused significant loss of basic protein without appreciably affecting MAG. Neither trypsin nor plasmin caused a specific cleavage of MAG to a derivative of MAG (dMAG) in a manner analogous to the endogenous neutral protease. Thus the endogenous protease appears unique in converting human MAG to dMAG much more rapidly than it degrades basic protein. MAG is slowly degraded along with other proteins when myelin is treated with trypsin or plasmin, but it is less susceptible to their action than is basic protein.     

Metabolism of Functional Groups Modifying the CNS Myelin-Associated Glycoprotein

We have examined the metabolic turnover of the peptide backbone of the CNS myelin-associated glycoprotein (MAG) and of the fucose and sulfate groups modifying this protein. Rats (20 or 90 days old) were injected intracranially with mixtures of [3H]fucose and [14C]glycine, [3H]glycine and [35S]sulfuric acid, or [3H]fucose and [35S]sulfuric acid. At times ranging from 30 min to 4 weeks later, myelin was isolated, and radioactivity in MAG was determined following electrophoretic separation. Following the peak of incorporation, glycine-derived radioactivity in the MAG peptide backbone declined several-fold during the first week and was then metabolically stable (half-life much greater than 1 month). Declines with time in [3H]fucose- and [35S]sulfate-derived radioactivity in MAG were similar to that of [3H]glycine, an observation indicating that the fucose and sulfate groups modifying MAG are metabolized together with the peptide backbone as a single metabolic entity. These results were confirmed by experiments involving selective immunoprecipitation of MAG. The rates of incorporation of labeled glycine, fucose, and sulfate into MAG all decreased approximately 12-fold between 20 days of age and adulthood, a finding providing further evidence for concerted turnover of the entire molecule. Because of this concerted turnover, we suggest that functional groups modifying MAG serve some permanent structural role in protein configuration.     

Myelin-Associated Glycoprotein: Electron Microscopic Immunocytochemical Localization in Compact Developing and Adult Central Nervous System Myelin

Light microscopic immunocytochemical studies have shown that myelin-associated glycoprotein (MAG) is localized in myelin of the developing CNS; but in the adult, MAG appears to be restricted to periaxonal regions of myelinated fibers. To extend these observations, we embedded optic nerves of 15-day-old rats, adult rats, and an adult human in epon after aldehyde and osmium tetroxide fixation. After 5% H2O2 pretreatment, thin sections were immunostained with 1:250-1:5,000 rabbit antiserum to rat CNS MAG according to the avidin-biotin-peroxidase complex (ABC) method. Dense deposits of reaction product covered compact myelin in both developing and adult optic nerves. When we used 1:500, 1:1,000, and 1:2,000 anti-MAG, less intense immunostaining of myelin was found. We also obtained the same localization in compact myelin with the peroxidase-antiperoxidase (PAP) method. With 1:250 anti-MAG, dense deposits of reaction product were not observed on axolemmal membranes or on oligodendroglial membranes located periaxonally and paranodally. In thin sections of adult human optic nerve, anti-MAG also stained compact myelin intensely. When thin sections of rat and human optic nerves were treated with preimmune or absorbed serum, no immunostaining was observed. Immunoblot tests showed that our MAG antisera did not react with any non-MAG myelin proteins. In contrast with earlier light microscopic data, this study shows that MAG localization does not change during CNS development; both developing and adult compact myelin sheaths contain MAG. As many biochemical studies also show that MAG is present in compact myelin, we suggest that this 100,000 dalton glycoprotein now be called myelin glycoprotein (MGP) instead of MAG.     

Calcium-Dependent Interaction Between the Large Myelin-Associated Glycoprotein and S100β

The myelin-associated glycoprotein is a transmembrane cell adhesion molecule expressed by myelinating glial cells of the nervous system. So far, only protein kinases have been reported to interact with the cytoplasmic domains of the two isoforms of the myelin-associated glycoprotein. We report here the identification of the first nonkinase intracellular ligand for the large isoform of the myelin-associated glycoprotein as the S100beta protein. The interaction is dependent on the presence of calcium. We have also localized the S100beta-binding site in the cytoplasmic domain specific to the large myelin-associated glycoprotein isoform to a putative basic amphipathic alpha-helix. A synthetic peptide corresponding to this region bound to S100beta in a calcium-dependent manner with a stoichiometric ratio of 1:1 (K(D) approximately 7 microM). We suggest that the observed interaction may play a role in the regulation of the myelinating glial cell cytoskeleton and the divalent cation-dependent signal transduction events during myelin formation and maintenance.     

Quantitation of the Myelin-Associated Glycoprotein in Human Nervous Tissue from Controls and Multiple Sclerosis Patients

Myelin-associated glycoprotein (MAG) was measured by radioimmunoassay in the human CNS and peripheral nervous system (PNS). The level of MAG, expressed as ng/microgram of total protein, was approximately 20-fold higher in whole homogenates of cerebral white matter (4.7 +/- 0.60) than of peripheral nerve (0.12-0.28). MAG concentrations were only slightly higher in the isolated myelin fractions from these tissues: CNS myelin, 5.6 ng/microgram; PNS myelin, 0.37 ng/microgram. The levels of MAG were measured in nine plaques, periplaque regions, and areas of macroscopically normal-appearing white matter (NAWM) from six separate multiple sclerosis brains and compared with the levels of other myelin proteins in the same samples. MAG and other myelin proteins were reduced to very low levels in plaques. The levels of MAG and basic protein (BP) and the activity of 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP) in periplaque areas were significantly lower than those in control white matter, and MAG and BP levels were also significantly reduced in NAWM. In a periplaque region and NAWM from the most rapidly progressing case of multiple sclerosis examined, the MAG content was between 30 and 35% of the control level, whereas BP and PLP levels and CNP activity were between 50 and 85% of control values. The reduction of MAG content in periplaque regions from all nine multiple sclerosis plaques examined was significantly greater than the reductions of BP level and CNP activity. In NAWM samples, the mean reduction of MAG content was also greater than the reductions of BP level and CNP activity, but the difference was only statistically significant in comparison to CNP.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cleavage of Myelin Basic Protein by Neutral Protease Activity of Human White Matter and Myelin

Polypeptides arising from neutral in vitro proteolysis of myelin basic protein (MBP) of human brain were evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. At pH 7 a marked breakdown of MBP resulted in the formation of 8-12 polypeptides ranging from 6 to 17 kd in molecular weight. As neutral proteolytic activity was not eliminated by either gel filtration or cation-exchange chromatography acid-soluble protease(s) involved probably have a size and electric charge similar to that of MBP. The enzymatic nature of neutral proteolysis was ascertained by heat inactivation and inhibition by alpha 2-macroglobulin. Incomplete inhibition of proteolysis and the failure of small peptides (less than 6 kd) to show up on electrophoresis seem to suggest that MBP was degraded by exopeptic proteases as well. Acid extracts of purified myelin yielded polypeptides similar to those of MBP of delipidated white matter. The results are consistent with a sequential limited proteolysis of MBP by neutral proteases probably associated with myelin and possibly related to the in situ catabolism of MBP in man.     

Sequential Limited Proteolysis of Myelin Basic Protein by Neutral Protease Activities of Bovine Brain

Acid extracts of delipidated white matter of bovine brain were prepared, and their proteolytic activities toward myelin basic protein (MBP) were evaluated at pH 3 and pH 7. This was done by measuring changes in total protein using a selective dye-binding assay, and by evaluating peptide patterns by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and densitometry. At pH 7 greater than 50% of total protein and about 75% of MBP were degraded after 48 h, whereas at pH 3 it was less than 20% altogether. Neutral proteolysis of MBP entailed up to 12 different proteolytic peptide fragments in the molecular weight range of 17.5 to 6 kd. Its enzymatic nature was verified using protease inhibitors, including N-ethylmaleimide, phenylmethylsulfonyl fluoride, o-phenanthroline, and EDTA, as well as pepstatin A and alpha 2-macroglobulin. Both transient changes in percentages of some intermediate peptides and differential effects of individual inhibitors on electrophoretic peptide patterns strongly suggest a sequential type of limited proteolysis. The results also indicate that acid extracts contained several endopeptidases of which a cysteine protease appears to initiate the breakdown of MBP.     

Electron Microscopic Immunocytochemical Localization of Myelin Proteolipid Protein and Myelin Basic Protein to Oligodendrocytes in Rat Brain During Myelination

Electron microscopic immunocytochemical studies were carried out to localize myelin basic protein and myelin proteolipid protein during the active period of myelination in the developing rat brain using antisera to purified rat brain myelin proteolipid protein and large basic protein. The anti-large basic protein serum was shown by the immunoblot technique to cross-react with all five forms of basic protein present in the myelin of 8-day-old rat brain. Basic protein was localized diffusely in oligodendrocytes and their processes at very early stages in myelination. The immunostaining for basic protein was not specifically associated with any subcellular structures or organelles. The ultrastructural localization of basic protein suggests that it may be involved in fusion of the cytoplasmic faces of the oligodendrocyte processes during compaction of myelin. Immunoreactivity in the oligodendrocyte and myelin due to proteolipid protein appeared at a later stage of myelination than did that due to basic protein. Staining for proteolipid protein in the oligodendrocyte was restricted to the membranes of the rough endoplasmic reticulum, the Golgi apparatus, and apparent Golgi vesicles. The early, uncompacted periaxonal wrappings of oligodendrocyte processes were well stained with antiserum to large basic protein whereas staining for proteolipid protein was visible only after the compaction of myelin sheaths had begun. Our evidence indicates that basic protein and proteolipid protein are processed differently by the oligodendrocytes with regard to their subcellular localization and their time of appearance in the developing myelin sheath.     

Jimpy Mice: Quantitation of Myelin-Associated Glycoprotein and Other Proteins

Myelin-associated glycoprotein (MAG), 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) activity, myelin basic protein (BP), and proteolipid protein (PLP) were quantitated in the brains of 20-day-old Jimpy and control mice. The levels of MAG, CNPase, and BP in Jimpy brains were 5.3%, 9.7%, and 1.9% of those in control brains, respectively. Immunoblotting analysis did not reveal an increased apparent Mr for MAG in the Jimpy mouse, as has been observed in some other hypomyelinating murine mutants. PLP was reduced more than the other proteins, as it was not detected by an immunoblotting technique that was capable of detecting 0.5% of the control level.     

Demyelinating Antibodies to Myelin Oligodendrocyte Glycoprotein and Galactocerebroside Induce Degradation of Myelin Basic Protein in Isolated Human Myelin

Abstract: Although the specificity of multiple sclerosis (MS) brain immunoglobulins (lgs) remains unknown, the incubation of these lgs with human myelin can lead to myelin basic protein (MBP) degradation mediated by neutral proteases. In this study, we demonstrate that monoclonal antibodies (mAbs) specific to myelin components such as the CNS-specific myelin oligodendrocyte glycoprotein (MOG) and galactocerebroside (GalC) are found to induce a significant loss of MBP mediated by neutral proteases in myelin. By contrast, antibodies to periaxonal and structural components of myelin, such as MBP and myelin-associated glycoprotein, are ineffective in inducing such MBP degradation. Among the 11 different anti-MOG mAbs directed to externally located epitopes of MOG, only two were found to induce a significant degradation of MBP, suggesting that antibody-induced MBP degradation is not only antigen specific but also epitope specific. Based on the inhibition of MBP degradation in the presence of EGTA and the analysis of the degradation products obtained following incubation of myelin with mAbs to GalC and MOG (8-18C5), the neutral protease involved in this antibody-induced degradation of MBP could be calcium-activated neutral protease. Taken together, these results suggest that antibodies to GalC and MOG can play a major role in destabilizing myelin through MBP breakdown mediated by neutral proteases and thus have an important role to play in the pathogenesis of MS.     

Endogenous Basic Protein Phosphatases in the Brain Myelin

Direct treatment of brain myelin with freezing/thawing in 0.2 M 2-mercaptoethanol stimulated the endogenous myelin phosphatase activity manyfold when 32P-labeled phosphorylase a was used as a substrate, a result indicating that an endogenous myelin phosphatase is a latent protein phosphatase. When myelin was treated with Triton X-100, this endogenous latent phosphatase activity was further stimulated 2.5-fold. Diethylaminoethyl-cellulose and Sephadex G-200 chromatography of solubilized myelin revealed a pronounced peak of protein phosphatase activity stimulated by freezing/thawing in 0.2 M 2-mercaptoethanol and with a molecular weight of 350,000, which is characteristic of latent phosphatase 2, as previously reported. Moreover, endogenous phosphorylation of myelin basic protein (MBP) in brain myelin was completely reversed by a homogeneous preparation of exogenous latent phosphatase 2. By contrast, under the same conditions, endogenous phosphorylation of brain myelin was entirely unaffected by ATP X Mg-dependent phosphatase and latent phosphatase 1, although both enzymes are potent MBP phosphatases. Together, these findings clearly indicate that a high-molecular-weight latent phosphatase, termed latent phosphatase 2, is the most predominant phosphatase responsible for dephosphorylation of brain myelin.     

Myelin-Associated Glycoprotein in the Central and Peripheral Nervous System of Quaking Mice

The myelin-associated glycoprotein (MAG) was quantitated in the CNS and PNS of quaking mice and the levels compared to the levels of myelin basic protein (MBP) and 2':3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) activity. In the brainstems of 36-day-old quaking mice, MBP, MAG, and CNPase were reduced to 12, 16, and 29% of control levels, respectively. In the sciatic nerves of the 36-day-old quaking mice, MBP and CNPase were 38 and 75% of control levels, respectively, whereas the concentration of MAG was unchanged or slightly increased. Similar quantitative results were obtained for the sciatic nerves and spinal roots of 7-month-old quaking mice. Immunoblots showed that the principal MAG band from the brainstems, sciatic nerves, and spinal roots of the quaking mice had a higher than normal apparent Mr. In addition, there was a minor component reacting with anti-MAG antiserum in the brainstems of the quaking mice that had a slightly lower Mr than control MAG and was not detected in the normal mice. The results for the quaking mice are compared with those from similar studies on other mutants with dysmyelination of the CNS and PNS.     

ADP-Ribosylation of Human Myelin Basic Protein

Abstract: When isolated myelin membranes were ADP-ribosylated by [³²P]NAD⁺ either in the absence of toxin (by the membrane ADP-ribosyltransferase) or in the presence of cholera toxin, the same proteins were ADP-ribosylated in both cases and myelin basic protein (MBP) was the major radioactive product. Therefore, cholera toxin was considered a good model for ADP-ribosylation of myelin proteins. Although purified human MBP migrates as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a molecular mass of 20 kDa, the microheterogeneity that is masked under these conditions can be clearly demonstrated on alkaline-urea gels at pH 10.6. At this pH, MBP is resolved into several components that differ one from the other by a single charge (charge isomers). These charge isomers can be resolved on CM52 columns at pH 10.6, and several can be ADP-ribosylated. Component 1 (C-1), the most cationic charge isomer, incorporated 1.79 mol of ADP-ribose/mol of protein. C-2 and C-3 (which differ from C-1 by the loss of one and two positive charges, respectively) incorporated slightly less at 1.67 and 1.63 mol of ADP-ribose/mol of protein, respectively, whereas C-8, the least cationic, incorporated less than 0.11 mol/mol of protein. In the presence of neutral hydroxylamine, the ADP-ribosyl bond was shown to have a half-life of about 80 min, suggesting an N-glycosidic linkage between ADP-ribose and an arginyl residue of the protein. As MBP contains several components that are ADP-ribosylated to different specific activities, the use of MBP, ADP-ribosylated in the natural membrane, to identify the sites involved would yield a mixture of peptides difficult to resolve. Therefore, to identify the sites ADP-ribosylated, an endoproteinase Lys-C digest of C-1 ADP-ribosylated by cholera toxin was prepared. Two radioactive peptides were isolated by reversed-phase HPLC. Amino acid and sequence analyses identified the radioactive peptides as residues 5–13 and 54–58 of the human sequence (sp. act., 0.89 and 0.62 nmol of ADP-ribose/nmol of peptide, respectively). The ADP-ribosylated residues were identified as Arg⁹ and Arg⁵⁴ by automated and manual Edman sequencing. Taken together with our previous observation that MBP binds GTP at a single site, these data suggest that MBP functions as part of a signal transduction system in myelin.     

Myelin-Associated Glycoprotein Is the Antigen for a Monoclonal IgM in Polyneuropathy

Recent studies show that IgM monoclonal antibody from patients with IgM paraproteinemia and peripheral neuropathy reacts with a protein component of human PNS myelin and an analogous component or components of human CNS myelin. We have now demonstrated that the antigen for this antibody is a specific glycoprotein component of myelin, referred to as myelin-associated glycoprotein (MAG). Human PNS and CNS myelin proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis on pore-gradient slabs, and MAG was identified by the immuno-electroblot procedure with rabbit anti-MAG (rat). The identical band(s) were stained by an analogous procedure with patient serum as the first antibody. Human PNS MAG had an apparent molecular weight of 107,000. Human CNS MAG appeared as three bands: 113,000, 107,000, and 92,000. Passage of myelin proteins through a concanavalin A-Sepharose column removed the staining component. Purified patient IgM, added to a lithium diiodosalicylate extract of myelin, immunoprecipitated MAG. This antibody also cross-reacted with MAG from bovine CNS, but not from rabbit, rat, or mouse.     

Production and Characterization of Monoclonal Antibodies Against Myelin-Associated Glycoprotein

Monoclonal antibodies against myelin-associated glycoprotein were generated by fusing mouse myeloma cells with spleen lymphocytes from BALB/c mice immunized with human myelin-associated glycoprotein purified from CNS myelin. Three groups of antibodies were identified: IgG antibodies recognizing the polypeptide moiety and IgG and IgM antibodies recognizing the carbohydrate moiety of the intact molecule. Properties of these antibodies were examined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the immunostaining technique using human CNS and peripheral nerve myelin, and ganglioside fractions isolated from human brain and peripheral nerve, and with immunohistochemical staining of human peripheral nerves. Part of human peripheral blood mononuclear cells was stained with the antibodies against the carbohydrate moiety, but not with IgG antibodies recognizing the polypeptide moiety. Natural killer activity was partially reduced after treatment of human peripheral blood lymphocytes with an IgM antibody and complement in vitro. The possibility that anti-myelin-associated glycoprotein antibodies might play a role in the pathogenesis of demyelinating diseases through modification of natural killer activity is discussed.     

Role of Myelin-Associated Neuraminidase in the Ganglioside Metabolism of Rat Brain Myelin

The role of myelin-associated neuraminidase in ganglioside metabolism was examined using rats of ages ranging from 17 to 97 days. The neuraminidase activity directed toward the ganglioside GM3 in the total myelin fraction was high during the period of active myelination and, thereafter, decreased rapidly to the adult level. The ganglioside composition became simpler during development with an increasing amount of GM1 and decreasing percentages of di- and polysialogangliosides. The decrease in the proportion of GD1a was most prominent, whereas relative amounts of GD1b and GT1b increased transiently before reducing to the adult levels. The heavy myelin subfraction contained higher percentages of di- and polysialo-species compared to the light myelin fraction at young and adult ages. The in vitro incubation of myelin of young rats under an optimal condition for neuraminidase action produced a profile of ganglioside changes similar to that observed in in vivo development. These results strongly suggest that myelin-associated neuraminidase may play a pivotal role in the developmental changes in the ganglioside composition of rat brain myelin.     

The Large Isoform of Myelin-Associated Glycoprotein Is Scarcely Expressed in the Quaking Mouse Brain

Two polypeptide isoforms of myelin-associated glycoprotein (MAG) with molecular masses of 72 and 67 kDa are produced by alternative splicing of the exon 12 portion. Our previous work has demonstrated that in the quaking mouse brain this alternative splicing is lacking and that the mRNA coding the large MAG isoform (L-MAG) is scarcely expressed, whereas that of small MAG isoform (S-MAG) is overexpressed. In the present study, we prepared antisera specific to the S-MAG and L-MAG amino acid residues, respectively. Immunoblots showed that the L-MAG band was scarcely detectable in the quaking mouse brain, whereas the S-MAG band had an apparently higher molecular mass than in the normal control. Our immunohistochemical study also showed that L-MAG was scarcely stained in the quaking mouse brain. These results seemed to reflect a reduction in content of L-MAG mRNA and abnormal glycosylation in the quaking mouse brain.     

Immunoblot Identification of 13.5 Kilodalton Myelin Basic Protein in Goldfish Brain Myelin

Myelin isolated from goldfish brain shows a multilamellar structure with a major dense line and two intraperiod lines. Sodium dodecyl sulfate gel electrophoresis revealed that the protein profile of goldfish brain myelin is distinctly different from that of rat brain myelin. No protein migrating to the position of proteolipid protein or DM-20 was seen in goldfish myelin. Goldfish acclimated to 5 degrees, 15 degrees, and 30 degrees C showed no qualitative differences in myelin proteins. The 13.5 kD protein in goldfish brain myelin and brain homogenate was intensely immunostained with the antiserum to human basic protein by the immunoblot technique. In contrast, none of the proteins of goldfish myelin were immunostained with antiproteolipid protein serum; however, both proteolipid protein and DM-20 of rat brain myelin were immunostained. The significance of the synthesis of myelin proteins by astrocytes in the goldfish brain is discussed.     

Myelin Basic Protein Is a Zinc-Binding Protein in Brain: Possible Role in Myelin Compaction

The zinc-binding proteins (ZnBPs) in porcine brain were characterized by the radioactive zinc-blot technique. Three ZnBPs of molecular weights about 53 kDa, 42 kDa, and 21 kDa were identified. The 53 kDa and 42 kDa ZnBPs were found in all subcellular fractions while the 21 kDa ZnBP was mainly associated with particulate fractions. This 21 kDa ZnBP was identified by internal protein sequence data as the myelin basic protein. Further characterization of its electrophoretic properties and cyanogen bromide cleavage pattern with the authentic protein confirmed its identity. The zinc binding properties of myelin basic protein are metal specific, concentration dependent and pH dependent. The zinc binding property is conferred by the histidine residues since modification of these residues by diethyl-pyrocarbonate would abolish this activity. Furthermore, zinc ion was found to potentiate myelin basic protein-induced phospholipid vesicle aggregation. It is likely that zinc plays an important role in myelin compaction by interacting with myelin basic protein.     

Myelin-Associated Glycoprotein Shares an Antigenic Determinant with a Glycoprotein of Human Melanoma Cells

A sulfated 100K-dalton glycoprotein has been shown to be released into the culture medium of melanoma cells. Monoclonal antibodies 10C5 and 11B5, which were raised to human melanoma cells, as well as HNK-1 bind to this glycoprotein. It is shown here that mouse anti-myelin-associated glycoprotein (MAG) carbohydrate antibodies raised to human MAG and a human IgM paraprotein associated with neuropathy also bind to the same 100K molecule. However, anti-MAG antibodies recognizing peptide epitopes do not appear to react with this glycoprotein of melanoma cells, a result suggesting that its similarity to MAG is restricted to shared carbohydrate moieties. The anti-melanoma antibodies (10C5 and 11B5) resemble HNK-1 in binding to MAG and to some 19-28K-dalton glycoproteins and sulfated, glucuronic acid-containing sphingoglycolipids of the peripheral nervous system (PNS). In addition, the anti-melanoma antibodies cross-react with neural cell adhesion molecule (N-CAM), an observation emphasizing the shared antigenicity between MAG and other adhesion molecules. The results demonstrate that the anti-melanoma antibodies fall into a class of monoclonal antibodies (including HNK-1, human IgM paraproteins associated with neuropathy, anti-human MAG antibodies, and L2 antibodies) that are characterized by reactivity against related carbohydrate determinants shared by human MAG, N-CAM, and several protein and lipid glycoconjugates of the PNS.