Glycoconjugate biochemistry: structure-function relationship
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Glycoconjugates are ubiquitously distributed in nature and have been im plicated in distinctly significant biological functions including energy resource, protection, lubrication, structural support, cell adhesion, molecular and cellular recognition, receptors for hormones and viruses. In this thesis studies on the following glycoconjugates are presented: bacterial, Micrococcus lysodeikticus, cell wall, simian cervical mucin, human pulmonary mucin, bovine gallbladder mucin, sperm surface glycoproteins and glycoconjugates of malarial parasite, Plasm odium falciparum . The information obtained on the structure of these complex carbohydrates utilising chemical, enzymatic, immunological and physical methods provided insight into the understanding, in particular, of structure-function relationship, degradation and biosynthesis of these macromolecules. Studies on model com pounds and analytical methods, all of which are vitally important tools in the study of glycoconjugates, are also described.The carbohydrate prosthetic group of M icrococcus lysodeikticus cell wall was shown to consist of a glycan moiety linked to the protein and an antigenic polysaccharide attached to the glycan m oiety of the peptidoglycan through a phosphodiester group. A variety of model com pounds were synthesised to establish the structure of the carbohydrate moiety as well as to study the kinetics of the acid hydrolysis of the phosphodiester group linked to muramic acid and to the reducing terminus of glucose. The study was performed on Micrococcus lysodeikticus cell wall polymer resistant to lysozyme, elaborating the structural requirement for stability to the enzyme. Furthermore, a water soluble polymer from the Micrococcus lysodeikticus cell wall was isolated and characterised, a novel observation. The study on this polymer suggested the possible deficiencies in the biosynthesis or possible autolysis of the cell wall polymer.A large number of model com pounds were chemically synthesised to identify the structural features of the cell wall peptidoglycans and those of the antigenic polysaccharide. In addition, several compounds were chemically synthesised to obtain the model com pounds necessary to conduct kinetic studies to define the type of linkage, i.e., differentiate between the monophosphate or pyrophosphate, between the cell wall polysaccharide and peptidoglycan, more precisely the linkage between muramic acid 6-phosphate of the peptidoglycan and the reducing terminal residue, glucose, of the polysaccharide.Comprehensive studies on simian cervical mucus glycoproteins were perform ed to relate the morphological and biophysical changes during the ovarian cycle to biochemical changes in the biochemical structure of the mucus. The biophysical and biochemical changes in the mucus are of vital significance in human reproductive physiology. The bonnet monkey produces larger amounts of mucus and its ovarian / menstrual cycle is similar to that of human. The extensive structural investigations on cervical glycoproteins of different phases of the ovarian cycle led to the elucidation of several interesting and novel structural features, such as the linkage of sialic acid to ultimate Nacetylgalactosoamine residues linked to serine or threonine of the protein core as well as to the galactose residues further away from the protein core. These two differentially linked sialic acid residues have been postulated to contribute to the function of mucus glycoproteins as well as to biophysical properties. Similarly, sulfate groups in the glycoprotein safeguard the integrity of the m acromolecule. Also, a novel type of a-galactosyl-linkage has been shown by us in the midcycle cervical mucus. In addition, it was also discovered by us that a-linked N-acetylgalactosoam ine residues were present in the midcycle cervical glycoproteins. The functional role of these a -linked residues is not clear. It is believed that this type of galactose residues are uncommon in man and old world monkey. The structural studies on luteal phase glycoproteins demonstrated significant differences in the structure of oligosaccharides. Similarities in the oligosaccharides structures were also observed. Polyclonal antibodies to midcycle bonnet monkey glycoproteins clearly demonstrated the functional role of the cervical mucus. It was discovered by us that the glycoproteins that line the cervical mucus channels are responsible for sperm migration. Our study, first of its nature, distinctly defined the functional role of the cervical glycoproteins and the significant contributions of the sugar residues. It was further shown that sperm penetration was inhibited by the antibody-mucus complex. The antibody against the midcycle glycoprotein cross-reacted with the midcycle mucus and weakly with the luteal phase mucus. Similarly the antibody against the luteal phase glycoprotein crossreacted with the luteal phase as well as, though weakly, with the midcycle mucus. These observations clearly suggested some common epitopes amongst glycoproteins of different phases of the ovarian cycle, interalia some common structural features in the glycoproteins of the different phases. The goal of this study, i.e., to relate the structure function relationship of the cervical mucus glycoproteins, was achieved. Initial work on sperm surface glycoproteins and gall bladder glycoprotein have been performed. Currently pursued research indicates that sperm surface glycoproteins may provide useful m eans to regulate fertility.Over a period of several years, extensive and detailed biosynthetic, biochemical and immunological studies were pursued to elaborate the role of the carbohydrate moiety of the variant glycoprotein antigens of the malarial parasite, Plasmodium falciparum . For many years it was known that sugars,such as N-acetylglucosamine, mannose and galactose w ere incorporated in several antigens including the vaccine candidates, i.e., M SP-1, 195 kDa glycoprotein, and MSP-2, 43-52 kDa glycoprotein. Labelled sugars were metabolically incorporated in MSP-1 and MSP-2. The labelled glycoproteins from the asexual erythrocytic stages of Plasmodium falciparum were purified to homogeneity by SDS-PAGE and the labelled sugars were identified. It was further observed that N -acetylglucosamine was incorporated in different glycoproteins in much more significant amounts than mannose, and mannose was in abundance compared to galactose. From these observations it was clear that N -acetylglucosamine was incorporated in glycoproteins in abundance in regions other than anchor as well. Our researches, for the first time, showed that N -acetylglucosamine was the sugar residue that provided the O-glycosyl type linkage to serine or threonine. Thus, the linkage point between the sugar moiety and the protein-core was defined. The studies on the carbohydrate moiety of these glycoproteins have become significantly important as these may provide clues to the immune evasion properties of the variant lycoprotein antigens.Reviews on secretory glycoproteins have been published. A review by us on malarial glycoprotein is in press. More recently a research article on the integration of glycans utilising glycophosphatidylinositol into plasma membranes and their possible role and interaction with the targeted cells has been discussed.Published Work: | p.4 | Synthesis of the 3- and 4-methyl, 3,4-dimethyl, and 3,4,6-trim ethyl ethers of methyl 2-acetamido-2-deoxy-a-D -m annopyranoside.Nasir-ud-D in and Rodger W. Jeanloz. Carbohydr. Res. (1973) 28, 243-251. | p.13 | Synthesis of the 6-methyl and 3,6-and 4,6-dimethyl ethers of methyl 2- acetam ido-2-deoxy-a-D -mannopyranoside. Nasir-ud-Din, Dorothy A. Jeanloz and Roger W. Jeanloz. Carbohydr. Res.( 1974) 38, 205-216. | p.25 | The chemical structure of a fragment of Micrococcus lysodeikticus cellwall. Nasir-ud-Din and Roger W. Jeanloz. Carbohydr. Res. (1976) 47, 245-260. | p.41 | Further purification and characterization of a circulating antigen in schistoso-iasis. T.E. Nash, Nasir-ud-Din and R.W. Jeanloz. J. Immunol. (1977) 119, 1627-1633. | p.48 | Amino- sugar phosphates from the cell wall of Micrococcus lysodeikticus. Nasir-ud- Din, Masashi Tomoda and Roger W. Jeanloz. Carbohydr. Res. (1977) 57, C1-C3. | p.51 | Studies of a carbohydrate-containing polymer from Cordia myxa. M.K. Bhatty, D.H. Shah, M.A. Saeed and Nasir-ud-Din. Pak.J. Sci. Res. (1978) 21, 162-163. | p.53 | Glycoproteins from Ustilago tritici. Nasir-ud-Din, J.K.W old, I. Matsumato, M I D. Chughtai and E. Walker-Nasir. Phytochem istry (1978) 17, 2059- 260. | p.55 | Synthesis of methyl (or propyl) 2-acetamido-2-deoxy D -glucopyran-oside 6-(a-D -glucopyranosyl phosphate) and derivatives for the study of the phosphoric ester linkage in Micrococcus lysodeikticus cell wall. Christopher D. Warren, Nasir-ud-Din and Roger W. Jeanloz. Carbohydr. Res. (1978) 64, 43-56. | p.69 | The chemical structure of a glycoprotein from the cervical mucus 89 (premenstrual) of Macca radiata. Nasir-ud-Din, R oger W. Jeanloz, Varnon N. Reinhold, James D. Moore and Janet W. M cArthur. Glycoconjugate Research (1979); J. Gregory and R.W. Jeanloz (eds), Academ ic Press, New York, Vol. 1, 241- 244. | p.73 | Role of synthetic phosphate diesters in study of bacterial cell wall. 73 Christopher D. Warren. Nasir-ud-Din, Vernon N. Reinhold and Roger W. Jeanloz. Glycoconjugate Research (1979) J. Gregory and R.W. Jeanloz (eds), Academic Press, New York, Vol. II, 839-841. | p.76 | Immunologically induced alteration in the morphology of the cervical 76 mucus of Macaca radiata. N a sir-ud-Din, Theodore E. Nash , Roger W. Jeanloz, Janet W. M cArthur and Dianne M. Gminski. Fertil. Steril. (1979) 32, 230-232. | p.79 | Cervical mucus glycoproteins in reproduction: Study of cervical mucus 79 from Macaca radiata. Nasir-ud-Din, Roger W. Jeanloz, Theodore E. Nash and Janet W. McArthur. Glycoconjugate. (1979), R. S chauer et al.,(eds.) George Theime, Stuttgart, 548-549. | p.81 | Changes in the cervical mucus of the bonnet monkey (Macaca radiata) during the menstrual cycle. Nasir-ud-Din, Janet W. McArthur and Roger W. Jeanloz. Animal Models for Research on Contraception and Fertility. (1979). Nancy J. Alexander (ed.) Harper and Row, Hargerstown, 396-403. | p.89 | Changes in the glycoprotein structure of the cervical mucus of the bonnet monkey during the menstrual cycle: Study of the premenstrual-phase mucus. Nasir-ud-Din, Roger W. Jeanloz, Vernon N. Reinhold and Janet W. McArthur. Carbohydr. Res. (1979) 75, 349-356. | p.98 | Isolation and purification of sperm atozoon-surface glcoprotein from Macaca radiatata. Nasir-ud-Din, Evelyne Walker-Nasir, Roger W. Jeanloz and Moshe Shalev. Carbohydr. Res. (1980) 85, C7-C9. | p.101 | Studies on bonnet monkey cervical mucus: The effect of proteases on mucus glcoproteins of Macaca radiata. Nasir-ud-Din, Roger W. Jeanloz, John R. Vercelotti and Janet W. McArthur. Biochim. Biophys. Acta (1981) 678, 483-496. | p.115 | Bonnet monkey cervical mucus glycoproteins: Study of the minor glycoprotein components of the periovulatory phase mucus. Nasir-ud-Din, Roger W. Jealoz and Janet W. M cArthur. Gtycoconjugates (1981) T. Yamakawa, T. Osawa and S. Handa (eds), Japan Scientific Societies Press, Tokyo, 328-329. | p.117 | Immunologically induced changes in macaque cervical mucus function: Inhibition of sperm penetration. Nasir-ud-Din Evelyne Walker-Nasir, Janet W. M cArthur, Theodore E. Nash, Moshe Shalev and Roger W. Jeanloz . Fertil. Steril. (1982) 37, 4 3 1 -4 3 5 . | p.122 | Isolation and characterization of bonnet monkey sperm surface qlycoproteins. Moshe Shalev, Nasir-ud-Din and Anwar Johar. Fed. Proc. (1 9 8 2 )4 1 . | p.123 | Fractionation of oligosaccharides containing sialic acid by liquid chromato-graphy on amino silica gel. G eneveive Lamblin, Andre Klein, A rnold Boersma, Nasir-ud-Din and Philppe Roussel. Carbohydr. Res. (1983) 118, C1-C4. | p.127 | Bonnet monkey cervical mucus glycoproteins: Study of the minor glcoprotein components of periovulatory phase mucus. Nasir-ud-Din, Roger W. Jeanloz, Philippe Roussel and Janet W. M cArthur. Biol. Reprod. (1983) 28, 1189-1199. | p.138 | Bonnet monkey cervical mucus: Isolation and charaterization of oligosacchaides from the Pronase-treated periovulatory phase glycoprotein. Nasir-ud-Din, E. W alker-N asir and M.A.K. Malghani. G lycoconjugates (1983) M .A.Chester et. al., (eds) Rahms, Lund, 603- 604. | p.140 | Role of epithelial secretions: G lycoproteins structure-function relationship. Nasir-ud-Din, M.A.K. M alghani and M.S. Ajaz. Proc. Poly. Sci. Sym. (1983) 2, 75-80. | p.146 | Modern trends: Glycoproteins. Nasir-ud-Din and E. Walker-Nasir. J. Univ. Bal. (1984) III, 1-11. | p.157 | The chemical structure of a high molecular weight fragment of Micrcoccus lysodeikticus cell wall. Nasir-ud-Din, E. W alker-N asir, S.AItaf Hussain, M.A.K. M alghani, T. Zamir and S.M. Ajaz. J. Univ. Bal. (1984) III, 55-72. | p.175 | Study of aspargine-linked glycopeptide derivatives by field-desorption mass spectrometry. Evelyne W alker-Nasir and Nasir-ud-Din. J. Univ. Bal. (1984) III, 47-54. | p.183 | The phosphate diester linkage of peptidoglycan polysaccharide moieties of Micrococcus lysodeikticus cell wall. Nasir-ud-Din, Michel Lhermitte, Geneviene Lamblin and Roger W. Jeanloz. J. Biol. Chem. (1985) 260, 9981-9987. | p.190 | Isolation, purification and partial characterization of a glycoprotein from plasma secretion. Nasir-ud-Din, M.A.K. Malghani and M.S. Ajaz. J. Chem. Soc. Pak. (1985) 7, 179-183. | p.195 | Glycoproteins in reprodution: Sperm surface and seminal plasma glycoproteins. Nasir-ud-Din. J. Univ. Bal. (1885) IV, 89-105. | p.212 | Glycoprotein structure-function relationship: Study of epithelial cell secretion. Nasir-ud-Din, M. Saleh Ajaz, and Irfanullah Khan. Natural Product Chemistry (1986) Atta-ur-Rahm an (ed.) Springer-Verlag, Berlin, 281-306. | p.238 | Isolation and partial characterization of human gallbladder mucin glycoprotein. Nasir-ud-Din, M.A.K. M alghani, M. Saleh Ajaz, S. Altaf Hussain and Abdullah Jan. G lycoconjugates (1985) E.A. Davidson et.al., (eds.) Praeger, New York, 82-83. | p.240 | Structure of sialyloligosaccharides isolated from bonnet monkey (Macaca radiata) cervical mucus exhibiting multiple blood group activities. Nasir-ud-Din, Roger W. Jeanloz, Genevieve Lamblin, Philippe Roussel, Herman van Hallbeek, Johanna H.G.M. Mutsaers and Johannes F.G. Vliegenthart. J. Biol.Chem. (1986 ) 261, 1992-1997. | p.247 | Epithelial secretions: Mucus glycoproteins. Nasir-ud-Din, Essays on Science (1986) H.M. Saeed (ed.) Hamdard Foundation Press, Karachi, Pakistan. 247-263. | p.264 | Localization of sulphate groups in a glycoprotein from hum an bronchial mucus of patients suffering from chronic bronchitis. Nasir-ud-Din, M.A.K. Malghani, M.Kasi, M. Lhermitte, P. Roussel and Evelyne Walker- Nasir. J. Chem. Soc. Pak. (1986) 8, 201 -2 08. | p.272 | Structure of acidic oligosaccharides isolated from Pronase-treated glycoprotein of bonnet-monkey (Macaca radiata) cervical mucus. Nasir-ud-Din. Carbohydr. Res. (1987) 159, 95-107. | p.285 | Glycoproteins: Biologically active macromolecules. Nasir-ud-Din, Evelyne W alker-N asir, M.A.K. Malghani anf S. A ltaf Hussain. J. Chem. Soc. Pak. (1988) 10, 471-481. | p.296 | Isolation, purification and partial characterization of neutral oligosaccharides from bovine gallbladder mucin glucoprotein. Nasir-ud-Din, M .Saleh Ajaz, S.Altaf Hussain, Bernard F. Smith and J. Thom asam ont. J. Chem. Soc. Pak. (1988) 10, 11-18. | p.304 | Partial charaterization of neutral oligosaccharides from normal human brochial secretion. Nasir-ud-Din, Evelyne Walker-Nasir, Mahmood Raza and Abdul M alik Kasi. J. Chem. Soc. Pak. (1988) 10, 33-41. | p.313 | Chemical structure of shark cartialge chondrointin sulfate. Anund Hallen. Nasir-ud-Din and Roger W. Jeanloz. J. Chem. Soc. Pak.(1989) 11, 168-177. | p.323 | Carbohydrtae moiety of Plasmodium falciparum . Nasir-ud-Din, R. Drager-Dayal, C. Decrind, D C. Hoessli, M.H. Qazi, G. Del Guidice and P H. Lambert. J. Chem. Soc. Pak. (1990) 12, 344-350. | p.330 | Studies of cervical glycoproteins: Isolation and characterization of neutral oligosaccharides from Pronase-treated glycoproteins of bonnet monkey (Macca radiata). Nasir-ud-Din, S. A ltaf Hussain, Roger W. Jeanloz and Evelyne Walker-Nasir. Carbohydr. Res. (1990) 205, 444- 452. | p.339 | Presence of O-glycosylated glycoproteins in the Plasmodium falciparum parasite. R. Dayal-Drager, Daniel C.Hoessli, C hristine Decrind, G uiseppe Del Guidice, Paul-Henri Lambert and Nasir-ud-Din. Carbohydr. Res. (1991) 209, C5-C8. | p.343 | Plasmodium falciparum synthesizes O-glycosylated glycoproteins containing O-linked N-acetylglucosam ine. Nasir-ur-Din, Renu Drager-Dayal, C hristine Decrind, Bei-Hong Hu, Giuseppe Del Guidice and Daniel Hoessli. Biochem. Int. (1992) 27, 55-64. A water soluble peptidoglycan-containing polymer from the Micrococcus lysodeikticus cell wall. Nasir-ud-Din, Fayyaz-ud-D in and Evelyne Walker-Nasir. Biochem. Soc. Trans. (1992) 20, 388S. | p.343 | Plasmodium falciparum synthesises O -glycosylated glycoproteins containing O -linked N-acetylglucosam ine. Nasir-ur-Din, Renu Drager- Dayal, C hristine Decrind, Bei-Hong Hu, G iuseppe Del G uidice and Daniel Hoessli. Biochem. Int. (1992) 27, 55-64. | p.353 | A water soluble peptidoglycan-containing polym er from the M icrococcus lysodeikticus cell wall. Nasir-ud-Din, Fayyaz-ud-D in and Evelyne Walker-Nasir. Biochem. Soc. Trans. (1992) 20, 388S. | p.354 | Plasmodium falciparum synthesises 43000 daltons protein containing O-linked glucosam ine. Nasir-ud-Din, M. Hassan, M.H. Qazi, Fayyaz-ud-Din, G. Sinaldi, D. Hoessli and E. Walker-N asir. Biochem. Soc. Trans. (1992) 20, 388S. | p.355 | Structure of a Micrococcus lysodeikticus cell wall fragment containing phosphorylated sugars. M. Siddiqui, M.H. Qureshi, M.A.K. M alghani, . Walker-Nasir, ayyaz-ud-D in and Nasir-ud-Din. iochem. Int. (1992) 26, 509-519 | p.366 | Further studies on cervical glycoproteins: Isolation and characterization of oligosaccharides from follicular phase cervical glycopproteins of bonnet monkey. Nasir-ud-Din, Fayyaz-ud-D in and E. W alker-Nasir. Biochem. Soc. Trans. (1993) 21, 186S. | p.367 | The chemical structure of a high molecular weight fragment of Micrococcus lysodeikticuss cell wall. Nasir-ud-Din, E. Walker-Nasir, Zahid Mahmood and Fayyaz-ud-Din. J. Chem. Soc. Pak. (1993) 15, 71- 77. | p.374 | Further studies on Plasmodium falciparum glycoproteins: Charact-rition of 195 kDa glycoprotein. M.H. Qureshi, M.H. Qazi, G. Senaldi, D.C. Hoessli and Nasir-ud-Din. Biochem. Soc. Trans. (1993) 21, 185S. | p.375 | A water-soluble fragment of Micrococcus lysodeiticus cell wall, Nasir-ud-Din, R.W. Jealnloz, E. Walker-Nasir, Fayyaz-ud-Din, Zahid Mahmood, K. Mahmood, A.H. Khan and F.S. Rahmani. Biochem. Mol. Biol. Int. (1994) 32, 1129-37. | p.384 | Galactose residues in Plasmodium falciparum glycoprotein: Incorporation and elimination. M.A. Qazi, G. Senaldi F. Tacchini-Cottier, M.H. Qazi, Fayyaz-ud-D in, D C. Hoessli, E. W alker-Nasir and Nasir-ud-Din. Biochem. Soc. Trans. (1994) 22, 362S. | p.385 | Sialytransferases of the bonnet monkey cervical epithelium : Isolation and characterization. Nasir-ud-Din, E. Walker-Nasir, Fayyaz-ud-D in, Z. Mahmood and A.H. Khan. Biochem. Soc. Trans. (1994) 22, 363S. | p.386 | Role of Cell surface carbohydrates in malaria. Nasir-ud-Din, M.A. Qazi, G. Senaldi, A.H. Khan, K. Mahmood, Z. Mahmood, M.H. Qazi, D.C. Hoessli and E. Walker-Nasir. Pure & Appl. Chem. (1994) 66, 2259-2262. | p.390 | Integration of mycobacterial lipoarabinom annans into glycosyphosphatidylinositol- rich domains of lymphomonocytic cell plasma membranes. Subburaj llangum aran, Stephan Am i, Monique Poincelat, Jean-Marc Theler, Patrik J. Brennan, Nasir-ud-Din and Daniel C. Hoessli. J. Immunol. (1995) 155, 1334-1342.Unpublished Work: | p.399 | Structural strudies on sialylated oligosaccharides of bonnet monkey (Macaca radiata) luteal phase cervical mucus. Z. Mahm ood, K. Mahmood, Abbas H. Khan, Javed Gondal, Evelyne Walker-Nasir, Daniel C. Hoessli, Elisabeth Rungger-Braendle, Roger W. Jeanloz and Nasirud-Din. | p.428 | Carbohydrate moiety of Plasmodium falciparum glycoproteins: the nature of the carbohydrate-peptide linkage in the 42-48 kDa glycoprotein. A.H. Qazi, M.H. Qazi, D.C. Hoessli, G. Senaldi, A.H. Khan, J. Gondal, E. Walker-Nasir and Nasir-ud-Din. | p.458 | Glycobiology of Plasmodium falciparum', an emerging area of research. D.C. Hoessli, E.A. Davidson, R.T. Schwarz and Nasir-ud-Din.