IUB commission of editors of biochemical journals the citation of bibliographic references in biochemical journals recommendations (1971)

A pentose-rich acidic glycoprotein was isolated from protease digested bovine vitreous humor by fractionation on an AG1-X2 column using NaCl solution gradient.The material eluted at 0.35 M NaCl (glycoprotein) was electrophoretically heterogeneous at pH 8.6 after partial purification on Sephadex G-25. Gel filtration on G-100 resolved the glycoprotein into two fractions. These fractions differ in molecular weight; mol. wt approx. 95 000 material consisted of two components on electrophoresis and mol. wt approx. 28 000 material showed only a single component on electrophoresis. The lower molecular weight component was re-chromatographed on Sephadex G-100 yielding a single orcinol positive component which gave a homogeneous band on gel electrophoresis.Quantitative analysis of this material gave 30% protein, 7.0% pentose, 18.7% glucosamine, 9.2% galactosamine, 10.9% hexuronic acid and 16.1% hexose.Treatment with 0.5 M NaOH at 20°C for 24 h resulted in a 50% decrease in the threonine content suggesting the possible involvement of this amino acid in the protein-carbohydrate linkage group.Paper chromatography of the fraction hydrolysate demonstrated the presence of glucurone, xylose, arabinose, glucose and galactose.     

Structural studies on a glycoprotein isolated from alveoli of patients with alveolar proteinosis.

A major glycoprotein 36 000 molecular weight) has been isolated from lung lavage of patients with alveolar proteinosis and found to contain five residues of hydroxyproline, fifty residues of glycine, three residues of methionine, 3 mol of sialic acid, 4.4 mol of mannose, 4.0 mol of galactose, 6.0 mol of glucosamine, and 1 mol of fucose. Cyanogen bromide (CNBr) treatment of the glycoprotein resulted, as expected, in four peptides of apparent molecular weights of 18 000, 12 000, 5000 and 1000, respectively. The chemical compositions of the CNBr peptides indicate the presence of hydroxyproline and high amounts of glycine in all but one of the peptides; two of the four CNBr peptides contain carbohydrate. Gel filtration, acrylamide gel electrophoresis and end-group analyses of the native glycoprotein and its CNBr peptides indicate that the peptides are homogeneous. End-group analyses of the CNBr cleavage products assign the 18 000 molecular weight peptide to the NH2-terminal portion and the 1000 molecular weight peptide to the COOH-terminal portion of the native glycoprotein molecule. Pronase digestion of the 36 000 molecular weight glycoprotein, followed by gel filtration and cation exchange chromatography, resulted in two fractions. One fraction was acidic and contained all the carbohydrate, a high content of aspartic acid and no hydroxyproline. The other fraction was basic and contained 8.4% hydroxyproline, 14% proline, 28% glycine and no carbohydrate, suggesting the presence of collagen-like sequence in the peptide chain. Paper electrophoresis of the basic fraction demonstrated two components, the amino acid compositions of which are identical to those of collagen. Partial amino-terminal sequence analysis of one of the CNBr peptides (18 000 molecular weight) indicated the presence of -Fly-Pro-HyP-Gly-sequence in the peptide chain, which confirms our suggestion that collagen-like regions are present in the native glycoprotein molecule. Limited acid hydrolysis of the acidic fraction and subsequent fractionation of the acid hydrolysate using Dowex column yielded a fraction which produced brown colour with ninhydrin reagent. Paper chromatography of this fraction demonstrated a large component which also stained brown with ninhydrin reagent. After acid hydrolysis, this component was found to consist of equal amounts of asparitic acid and glucosamine, indicating that the N-acetylglucosamine of the oligosaccharides is linked to the asparagine residue of the peptide. No serine or threonine linkages are present.     

Purification and characterization of the epitectin from human laryngeal carcinoma cells

The purification and partial characterization of epitectin (previously called Ca antigen) from a human cancer cell line is described. This glycoprotein, which is expressed on a wide range of human tumors and certain specialized normal epithelia, can be detected using monoclonal antibodies, Ca1, Ca2, and Ca3. The purified glycoprotein had a high density (1.40 g/ml) on isopycnic centrifugation indicating a high carbohydrate content. The molecular mass of epitectin as determined by size-exclusion chromatography ranged from 1.0 to 1.5 x 10(6) daltons. However, the purified epitectin gave two bands of apparent molecular weight 390,000 and 350,000 on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The isoelectric points of epitectin and asialoepitectin were found to be 5.3-5.4 and 6.8, respectively. The oligosaccharides were isolated from metabolically labeled epitectin by alkaline borohydride treatment and their structures established based on high performance liquid chromatography and paper electrophoretic migration, sugar composition, the results of sequential exoglycosidase treatment, periodate oxidation, and methylation analysis. The structures of the three major fractions, which together account for about 80% of the radioactivity, were assigned as NeuNAc alpha 2----3Gal beta 1----(NeuNAc alpha 2----6)3GalNAc(OH), NeuNAc alpha 2----3Gal beta 1----3GalNAc(OH), and Gal beta 1----3 GalNAc(OH). The structures of the minor fractions were tentatively assigned as NeuNAc----Gal(NeuNAc----Gal----GlcNAc)----GalNAc(OH), Gal beta 1----(NeuNAc alpha 2----6)3GalNAc(OH), NeuNAc alpha 2----6GalNAc(OH), and GalNAc(OH). It is proposed that the protein sequence and/or the distribution of the saccharides on the protein core are the determinants on epitectin that are recognized by the Ca antibodies.     

Characterization of arabinose and ferulic acid rich pectic polysaccharides and hemicelluloses from sugar beet pulp

Pectic polysaccharides were extracted from sugar beet pulp to yield fractions representing homogalacturonans, rhamnogalacturonans, arabinans and relatively small amounts of glucomannans and xyloglucans. The homogalacturonans had an apparent molecular weight of 21 kDa and contained relatively high amounts of methyl esters and relatively low amounts of acetyl groups as compared with the ramified 'hairy' regions. Three populations which originated from the ramified 'hairy' regions of pectin were distinguished. Two of these were rhamnogalacturonans with high apparent molecular weights of 1300 and 120 kDa, respectively. These populations had a high Ara and ferulic acid content. Despite the high neutral sugar content, these rhamnogalacturonans strongly bound to a DEAE column. The third population which originated from the ramified 'hairy' regions was a neutral population, which did not interact with the DEAE column and had a low apparent molecular weight and a high Ara and ferulic acid content. The arabinan side-chains of the rhamnogalacturonans were heavily branched in all populations. Enzymatic degradation of the xyloglucans showed similarities with apple xyloglucans with respect to the substitution with Fuc and Gal.     

Purification and characterisation of alpha-L-fucosidase from human placenta. pH-dependent changes in molecular size.

alpha-L-Fucosidase has been purified 12 000 fold from human placenta. The enzyme is a glycoprotein containing, by weight: 0.9% galactose; 1.9% mannose, 1.9% N-acetylglucosamine and 1.9% N-acetylneuraminic acid. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate separated proteins with molecular weights ot 55 000, 51 400 and 25 000. Resolution of the two larger protein bands varied with the gel system and these proteins may differ only in carbohydrate content. Gel filtration of te purified enzyme failed to separate the three proteins. Treatments with the cross-linking reagent dimethyl suberimidate prior to electrophoresis, resulted in a diminution of the original protein bands and the formation of oligomers with molecular weights of 80 000, 100 000, 130 000, and 144 000. These results suggest that the heavy (55 000 and 51 400) and light (25 000) proteins are structurally associated. The molecular weight of the native enzyme, measured by gel filtration, was dependent on the pH of the eluting buffer. At pH 5.0 or 6.0 a catalytically active peak was observed, with a molecular weight of 305 000. At pH 7.5 this peak was completely absent and the enzyme eluted as an asymmetrical peak with an apparent molecular weight of about 60 000. The reduction in apparent molecular weight at pH 7.5 was reversible by dialysis of isolated fractions at pH 6.0. In agreement with these findings the sedimentation coefficient was 8.5 S at pH 5.0 but only 3.6 S at pH 7.5. The results can be accounted for by the existence of a pH-dependent equilibrium between aggregated and dissociated forms of the enzyme or by pH-depedent conformational changes.     

红栓菌多糖的分离纯化与分析

液体发酵得到的红栓菌（Ｐｙｃｎｏｐｏｒｕｓｃｉｎｎａｂａｒｉｕｓ）菌丝经沸水抽提,胰匀浆处理和Ｓｅｖａｇｅ法除蛋白,ＤＥＡＥＳｅｐｈａｄｅｘＡ－２５与ＳｅｐｈａｄｅｘＧ－１５０柱层析,乙醇沉淀得到二种多糖组分（简称ＰＣ＿１、ＰＣ＿２）。经聚丙烯酰胺电泳,冻融分析鉴定为均一体。ＰＣ＿１和ＰＣ＿２的,总糖含量分别为８１．５％和７７．３％,分子量分别为２３０００和１３０００．两者的红处光谱具多糖特征吸收峰,在紫外区无明显吸收现象,基本不含氮。ＰＣ＿１单糖组成为Ｄ－葡萄糖、Ｄ－半乳糖及Ｄ－甘露糖,ＰＣ＿２为Ｄ－半乳糖、Ｄ－甘露糖。     

Liver as a source of transformed cell growth factor

Isolated rat hepatocytes release an acidic glycoprotein(s) that can selectively promote the growth of transformed cells. This factor has a molecular weight of 60 000–70 000 D. Liver microsomal and cytosol fractions contain two species of stimulatory activity—44 000 and 3 500 D. Mitochondrial and nuclear fractions contain only the lower molecular weight factor.     

Studies on Stigma Pellicle Glycoproteins of Raphanus sativus L.

Stigma surface diffusates of Raphanus sativus were subjected to polyacrylamide gel electrophoresis. There appeared protein bands, one of which gave positive PAS reaction, indicating it was glycoprotein. SDS polyacrylamide gel electrophoresis showed that the stigma diffusates contained many protein bands. By comparing their mobilities with those of standard proteins of known molecular weights, the molecular weights of some of the major fractions were estimated to be 15000, 30000—46000 and 70000 daltons. After Schiff reagent staining two main glycoprotein fractions appeared on the SDS gel electrophoretic pattern. Their molecular weights were estimated to be lower than 15000 and higher than 100000 daltons. By using acrylamide gel isoelectric focusing method, it was found that the stigma surface diffusates contained an acidic glycoprotein with pH of about 3.7. The amino acid composition of the purified stigma glycoprotein was determined with amino acid analyzer. Glycine, glutamic acid, serine, aspartic acid were some of the predominant amino acids. The diffusates were analysed by gasliquid chromatography for sugars. Results showed that the carbohydrate fraction of the glycoprotein consisted of arabinose 17.3%; galactose 19.1%, xylose 8.1%, mannose 5.4%, glucose 23.7%, rhamnose and/or fucose 26.4%. In the stigma surface diffusates of Raphanus sativus, the content of protein was estimated to be 16% and that carbohydrate was 11%.     

Characterization of an oviductal glycoprotein associated with the ovulated hamster oocyte

Hamster oviducts in culture incorporate [35S]-methionine into secretory proteins. One of these proteins is immunoprecipitated by a monoclonal antibody specific to an antigen found in oviductal oocytes but not in ovarian oocytes. This antigen, called oviductin, is progressively added to the oocyte during its transit through the oviduct. Oviductin migrates as a diffuse band with a molecular mass between 160 and 250 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. The electrophoretic behavior of this protein suggests the presence of polysaccharide side chains. Chemical deglycosylation causes a decrease in molecular mass and removes the antigenic determinant originally present on the glycoprotein. By using the radiation inactivation method, the molecular mass of the core protein has been found to be approximately 44 kDa. These results indicate that the oviduct is an actual site of synthesis of the oviductin. This glycoprotein contains a high proportion of sugar residues, which account for antigenic determinant recognized by the monoclonal antibody.     

An analysis of the protein, glycoprotein and monosaccharide composition of Dictyostelium discoideum plasma membranes during development.

Qualitative and quantitative changes in the protein and glycoprotein components of the plasma membrane of the cellular slime mould Dictyostelium discoideum have been detected by analysis of sodium dodecyl sulphate-polyacrylamide gel electrophoretic patterns. The amounts of proteins of subunit molecular weight 220 000, 91 000, 63 000, 59 000, 56 000 increased during the acquisition of aggregation competence, while proteins of subunit molecular weight 82 000 and 22 000 decreased. The amounts of glycoproteins with apparent subunit molecular weights 285 000, 150 000, 137 000, 100 000, 53 000, 50 500 and 30 500 increased during differentiation while a 125 000 dalton component decreased dramatically in amount. The neutral and amino sugar composition of the plasma membrane was also analyzed and found to remain essentially unchanged during the first 12 h of differentiation. The major sugars were mannose, fucose, and glucosamine; galactose and galactosamine were also present, but in lower amounts.     

Isolation of human platelet glycoproteins.

Human platelet glycoproteins were isolated from whole platelets by two methods. The first method, that of affinity chromatography on wheat germ agglutinin, is based on the known affinity of lectins for cell surface glycoproteins. When solubilized whole platelets are used as starting material for this procedure, elution with N-acetylglucosamine yields primarily a glycoprotein of Mr approximately 150 000 as estimated by sodium dodecyl sulfate-acrylamide gel electrophoresis. The second method is based on the ability of the chaotropic salt lithium diiodosalicylate to extract glycoprotein from particulate cell fractions in water-soluble form. This method yields three major glycopeptides with apparent molecular weights after sulfhydryl reduction of 145 000, 125 000, and 95 000 as estimated on 5.6% sodium dodecyl sulfate-acrylamide gels. Carboxymethylation of these preparations in the presence of sulfhydryl-reducing agent further resolves a glycoprotein of Mr approximately 165 000. Treatment of whole platelets by periodate oxidation and sodium[3H]-borohydride reduction labels the three major glycoproteins extracted by lithium diiodosalicylate and the glycoprotein of Mr approximately 150 000 isolated on wheat germ agglutinin confirming their surface orientation. However, glycoprotein with Mr approximately 165 000 resolved by carboxymethylation of the lithium diiodosalicylate extracted glycoprotein mixture was not labelled by this method, suggesting that it represents the granule protein with similar electrophoretic characteristics described by others. Phosphorylation of intact platelets with 32Pi also results in labelling of glycoproteins isolated by both methods, suggesting that these molecules traverse the bilipid layer of the platelet membrane, bearing reactive groups on both outer and cytoplasmic surfaces.     

Purification and properties of the main coagulant and anticoagulant principles of Vipera russellii snake venom

Vipera russellii venom was separated into thirteen fractions by means of DEAE-Sephadex A-50 column chromatography. Fraction III possessed anticoagulant and phospholipase A activities and Fraction XI possessed procoagulant and caseinolytic activities, both were further purified by gel filtration on Sephacryl S-200 column. Purified procoagulant (Component II) was a two-chain protein with molecular weight of 86 000 consisting of A-chain (Mr 66 000) and B-chain (Mr 20 000). It was a glycoprotein containing 7.8% neutral sugar and 715 amino-acid residues. The procoagulant activity was 10-times that of the crude venom. It was an acidic proteinase with isoelectric point of pH 4.2. Upon heat treatment at 60 degrees C, Component II was stable at pH 5.5 and 7.2 for 3 h, but was destroyed completely after 30 min at pH 8.9. It was devoid of esterase or amidase activity. Purified anticoagulant (Component I) was a single peptide chain with molecular weight of 16 000. It was carbohydrate free and contained 136 amino-acid residues. It was a basic protein with an isoelectric point of larger than pH 10. It was a potent phospholipase A with an enzymatic activity of 510 +/- 30 mumol/min per mg using phosphatidylcholine as substrate, and 1 microgram/ml was sufficient to cause 100% hemolysis by the indirect hemolytic method. Upon heat treatment at 90 degrees C, Component I was heat stable at pH 5.5 for more than 3 h, but was destroyed completely after 2 h at pH 7.2 and 8.9. The anticoagulant activity of Component I could be neutralized by platelet factor 3, tissue thromboplastin and cephalin.     

Lymphocyte plasma membranes. VI. Plasma membrane glycoproteins of thymic and splenic lymphocytes from inbred rats.

Plasma membranes of splenic and thymic lymphocytes from ACI rats were analyzed for their protein and glycoprotein components by surface radioiodination with 125I and SDS-polyacrylamide gel electrophoresis. The glycoproteins were extracted with lithium diiodosalicylate, characterized and assayed with antisera to thymic antigen. Plasma membranes of both cell types showed more than 25 proteins of which 10--15 were glycoproteins. Both cells showed five major glycoproteins but their apparent molecular weights or intensities differed. Surface radioiodination showed a 120 000 daltons component, common to both cell types, and a 27 000 daltons thymus-specific component as the most exposed surface glycoproteins. Lithium diiodosalicylate extracts of the plasma membranes contained almost all of the glycoprotein components and comprised 5-6 percent of the total membrane protein and 40-50 percent of the total membrane carbohydrate, with sialic acid content in thymus twice that of the spleen cells. About 1 percent of the total plasma membrane protein and 7 percent of the total isolated glycoproteins from thymocytes were reactive with rabbit anti-rat thymocyte antiserum and the immune precipitates showed two components with apparent molecular weights of 72 000 and 27 000.     

Structure of egg yolk very low density lipoprotein. Polydispersity of the very low density lipoprotein and the role of lipovitellenin in the structure

Egg yolk very low density lipoprotein contained on the average 75% of lipid which could be extracted by ether and 25% of a residual lipoprotein, the classical lipovitellenin. The ether-extracted lipid was composed of 75% triglycerides, 7% sterols, 2% mono- and diglycerides, and 16% phospholipids. Lipovitellenin contained 48% lipid composed of 87% phospholipids, 11% triglycerides, and 2% sterols. The protein vitellenin was composed for the most part of units of 74,000 and 270,000 daltons molecular weight.Egg yolk very low density lipoprotein is polydisperse. Preparative ultracentrifugation separated it into six fractions of different average molecular size, and gel chromatography separated it into five. The fractions of larger molecular size contained more lipid and triglyceride than did the fractions of smaller molecular size. The proteins of the fractions appeared to be similar.Egg yolk very low density lipoproteins appear to be a series of molecules composed of cores of lipid of varying sizes with each core surrounded by a layer of lipovitellenin, which is composed principally of glycoprotein and phospholipid.     

Partial deglycosylation of blood-group-specific glycoproteins.

The time course for the partial deglycosylation of blood-group-specific glycoproteins from human ovarian-cyst fluids with 0.25 M-H2SO4/acetic acid and 6 M-HCl in methanol was studied. Either reagent readily removed about 80% of the carbohydrate from the glycoproteins to leave non-diffusible glycopeptides that contain N-acetylgalactosamine as the predominant sugar. Some changes in amino acid distribution were observed during the deglycosylation, which were attributed to an accelerated break-up of the nonglycosylated regions of the parent glycoprotein. The N-acetylgalactosaminyl-peptides isolated were judged to be polydisperse by gel filtration, and ion-exchange chromatography divided the glycopeptide population into several fractions with differing amino acid compositions. A Lumbricus terrestris hexosaminidase preparation was successful in removing almost all the remaining sugar from the glycopeptides, but caused further rupture of the peptide. When a per O-acetylated glycoprotein was treated with the H2SO4/acetic acid reagent the glycopeptide contained, in addition to N-acetylgalactosamine, about 50% of the sialic acid present in the parent glycoprotein, indicating that most of this sugar is located near the peptide end of the carbohydrate chains.     

Salient characteristics of bovine pituitary intraglandular colloid: a transport medium for determinants to pituitary hormones and immunoglobulins

Bovine hypophyseal intraglandular colloid, the holocrine secretion of marginal intermediate lobe cells, contains high molecular weight (MW 45 000 to 158 000) protein fractions and low molecular weight (MW below 25 000) peptide fractions. The fractions display immunoreactive determinants similar to those in pituitary hormones, cerebrospinal fluid (CSF), serum IgG and albumin. Immunoelectrophoresis shows that components in high molecular weight fractions distribute themselves in three distinct bands (gamma, beta, alpha), while those in low molecular weight fractions distribute themselves in two bands (gamma and alpha). Physiochemical characteristics, i.e., sedimentation rate, percentage of hexose, total CHO and 12C as well as the content of cystine are presented. It is concluded that pituitary colloid should not be dismissed as a waste product of cellular degeneration, since there are strong suggestions that it serves as a transport medium for certain pituitary hormones.     

Isolation and partial characterization of the major glycoproteins of horse and swine erythrocyte membranes

The major glycoproteins of horse and swine erythrocyte membranes were isolated and examined chemically and immunologically. The major glycoprotein of horse erythrocyte membranes had a molecular weight of 33 000 and consisted of 46.2% protein and 53.8% carbohydrate, of which 9.4% was hexose, 10.1% hexosamine and 33.7% sialic acid. This glycoprotein was associated with activity for the infectious mononucleosis heterophile antigen.There were two different major glycoproteins in swine erythrocyte membranes. One major glycoprotein had a molecular weight of 46 200 and consisted of 34.2% protein and 65.8% carbohydrate, of which 18% was hexose, 19% hexosamine and 27.2% sialic acid. This glycoprotein had phytohemagglutinin (Phaseolus vulgaris) binding activity. The other glycoprotein had a molecular weight of 29 000 and consisted of 50.4% protein and 49.6% carbohydrate, of which 6.4% was hexose, 7.0% hexosamine and 36.3% sialic acid. This glycoprotein had weak or absent phytohemagglutinin binding activity.     

Isolation and characterization of a hemagglutinin from Amphitrite ornata, a polychaetous annelid.

Extracts of the marine polychaetous annelid, Amphitrite ornata, agglutinate rat, rabbit, chicken and human erythrocytes and in other work have been shown to inhibit the growth of Ehrlich ascites tumors in mice. Fractionation of extracts on Sephadex G-100 gave three active fractions with molecular weights of 30 000, 54 000 and 100 000. The 30 000 dalton fraction (B) was purified 72-fold by ammonium sulfate precipitation, gel filtration and preparative disc gel electrophoresis. The purified hemagglutinin, amphitritin, was homogenous on analytical disc gel electrophoresis at four different pH values and gave a sharp boundary in sedimentation velocity ultracentrifugation. The three fractions showed paralled specificity toward rat and chicken erythrocytes, the former giving the higher titer. The purified agglutinin was active toward human blood groups A, B and O and exhibited 4-fold higher activity toward group A. The hemagglutinin titer against rat red blood cells was lowered only by N-acetylgalactosamine, the terminal sugar residue of the group A determinant. None of the saccharides tested inhibited agglutination of chicken erythrocytes. Hemagglutinin activity was insensitive to dialysis or treatment with EDTA. The activity was not affected by digestion with trypsin or pronase, but was destroyed by phenol extraction. Analytical disc gel electrophoresis showed one protein band with high anodal mobility at pH 8.5, which was not affected by proteolytic enzymes but was removed by phenol. Activity was unaffected by heating at 70 degrees C for 30 min but was destroyed by similar treatemtn at 85 degrees C. Activity was at a maximum at pH 7-9 and decreased reversibly down to pH 4 at which point it was irreversibly inactivated. The higher molecular weight agglutinin (A1) could be dissociated to give amphitritin by treatment with 6M urea of precipitation in 55% (NH4)2SO4. This dissociation was not reversed by dialysis. Amphitritin is a glycoprotein with a molecular weight determined by gel filtration of 30 000 and by approach to equilibrium sedimentation of 32 000. Amino acid analysis showed a preponderance of aspartic and glutamic acids and relatively large amounts of glycine, proline, alanine, valine and cysteine. The carbohydrate moeity which represented 12.8% of the molecule, contained mannose, galactose, glucosamine and sialic acid. Amphitritin is the first hemagglutinin to be isolated from a polychaetous annelid.     

Studies of a large transformation-increased membrane protein in the BHK21 cell system

We have recently described in BHK cells a plasma membrane protein of molecular weight 177 000, which is significantly increased in Hamster Sarcoma Virus-transformed cells (Lage-Davila, A. and Montagnier, L. (1977) Biochem. Biophys. Res. Commun. 79, 577–584). We present now a study of proteins from purified plasma membrane fractions in the same pair of clones. Solubilization conditions, cross-linking experiments, metabolic labelling and enzymatic radioiodination allow to characterize this 177 000 transformation-increased protein as an integral membrane glycoprotein partially exposed at the outer cell surface. Additional information on other membrane proteins in this system is also given.     

Facilitated glucose transporter of human erythrocyte: proteolytic mapping of the [3H]cytochalasin B photoaffinity-labeled transporter polypeptide

The human erythrocyte D-glucose transporter is an integral membrane glycoprotein with an heterogeneous molecular mass spanning a range 45-70 kDa. The protein structure of the transporter was investigated by photoaffinity labeling with [3H]cytochalasin B and fractionating the labeled transporter according to molecular mass by preparative SDS-polyacrylamide gel electrophoresis. Each fraction was digested with either papain or S. aureus V8 proteinase, and the labeled proteolytically derived peptide fragments were compared by SDS polyacrylamide gel electrophoresis. Papain digestion yielded two major peptide fragments, of approx. molecular mass 39 +/- 2 and 22 +/- 2 kDa; treatment with V8 proteinase resulted in two fragments, with mass of 24 +/- 2 and 15 +/- 2. Proteolysis of each transporter fraction produced the same pattern of labeled peptide fragments, irrespective of the molecular mass of the original fractions. The binding characteristics of [3H]cytochalasin-B-labeled transporter to Ricinis communis agglutinin lectin was examined for each transporter molecular mass fraction. It was found that higher-molecular-mass fractions of intact transporter had a 2-fold greater affinity for the lectin than lower-molecular-mass fractions (i.e., 67 kDa greater than 45 kDa fraction). However, proteolytically derived labeled peptide fragments from each fraction had minimal affinity for the lectin. These results suggest that the labeled peptide fragments have been separated from the glycosylated regions of the parent transporter protein. The present findings indicate that, although transporter proteins have an apparently heterogeneous molecular mass, some regions of the protein share a common peptide. Furthermore, the glycosylated regions appear to be located some distance from the [3H]cytochalasin-B-labeled site(s).