Abstract
Using primary hepatocytes in culture, various 2-acetamido-2-deoxy-D-glucose (GlcNAc) analogs were examined for their effects on the incorporation of D-[3H]glucosamine, [35S]sulfate, and L-[14C]leucine into cellular glycoconjugates. A series of acetylated GlcNAc analogs, namely methyl 2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-α-(3) and β-D-glucopyranoside (4) and 2-acetamido-1,3,4,6-tetra-O-acetyl-2-deoxy-D-glucopyranose (5), exhibited a concentration-dependent reduction of D-[3H]glucosamine, but not of [35S]sulfate incorporation into isolated glycosaminoglycans (GAGs), without affecting L-[14C]leucine incorporation into total protein synthesis. These results suggest that analogs 3–5 exhibit an inhibitory effect on D-[3H]glucosamine incorporation into isolated GAGs by diluting the specific activity of cellular D-[3H]glucosamine and by competing for the same metabolic pathways. In the case of the corresponding series of 4-deoxy-GlcNAc analogs, namely methyl 2-acetamido-3,6-di-O-acetyl-2,4-dideoxy-α-(6) and β-D-xylo-hexopyranoside (7) and 2-acetamido-1,3,6-tri-O-acetyl-2,4-dideoxy-D-xylo-hexopyranose (8), compound 8 at 1.0 mM exhibited the greatest reduction of D-[3H]glucosamine and [35S]sulfate incorporation into isolated GAGs, namely to ∼7% of controls, and a moderate inhibition of total protein synthesis, namely to 60% of controls. Exogenous uridine was able to restore the inhibition of total protein synthesis by compound 8 at 1.0 mM. Isolated GAGs from cultures treated with compound 8 were shown to be smaller in size (∼40 kDa) than for control cultures (∼77 kDa). These results suggest that the inhibitory effects of compound 8 on cellular GAG synthesis may be mediated by the incorporation of a 4-deoxy moiety into GAGs resulting in premature chain termination and/or by its serving as an enzymatic inhibitor of the normal sugar metabolites. The inhibition of total protein synthesis from cultures treated with compound 8 suggests a uridine trapping mechanism which would result in the depletion of UTP pools and cause the inhibition of total protein synthesis. A 1-deoxy-GlcNAc analog, namely 2-acetamido-3,4,6-tri-O-acetyl-1,5-anhydro-2-deoxy-D-glucitol (9), also exhibited a reduction in both D -[3H]glucosamine and [35S]sulfate incorporation into isolated GAGs by 19 and 57%, of the control cells, respectively, at 1.0 mM without affecting total protein synthesis. The inability of compound 9 to form a UDP-sugar and, hence, be incorporated into GAGs presents another metabolic route for the inhibition of cellular GAG synthesis. Potential metabolic routes for each analog's effects are presented.
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References
Poole AR, Biochem J 236, 1–14 (1986).
Hook M, Annu Rev Biochem 53, 847–69 (1984).
LeBaron RG, Hook A, Esko JD, Gay S, Hook M, J Biol Chem 264, 7950–6 (1989).
Gallagher JT, Lyon M, Steward WP, Biochem J 236, 313–25 (1986).
Ruoslahti E, Annu Rev Cell Biol 4, 229–55 (1988).
Keller KL, Keller JM, May JN, Biochemistry 19, 2529–36 (1980).
Esko JD, Rostand S, Weinke JL, Science 241, 1092–6 (1988).
Johnson KH, O'Brien TD, Betsholtz C, Westermark P, Lab Invest 66, 522–35 (1992).
Kjellen L, Bielefeld D, Höök M, Diabetes 32, 337–42 (1983).
WuDunn D, Spear PG, J Virol 63, 52–8 (1989).
Shieh M-T, WuDunn D, Montgomery RI, Esko JD, Spear PG, J Cell Biol 116, 1273–81 (1992).
Leong JM, Robbins D, Rosenfeld L, Lahiri B, Parveen N, Infec Immun 66, 6045–8 (1998).
Chen JCR, Stephens RS, Microb Pathog 22, 23–30 (1997).
Chen Q, Barragan A, Fernandez V, Sundström A, Schlichtherle M, Sahlén A, Carlson J, Datta S, Wahlgren M, J Exp Med 187, 15–23 (1998).
Coppel RL, Brown GV, Nussenzweig V, Curr Opin Microbiol 1, 472–81 (1998).
Sinnis P, Sim BKL, Trends Microbiol 5, 52–8 (1997).
Snow AD, Sekiguchi RT, Nochlin D, Fraser P, Kimata K, Mizutani A, Arai M, Schreier WA, Morgan DG, Neuron 12, 219–34 (1994).
Snow AD, Sekiguchi RT, Nochlin D, Kalaria D, Kimata K, Am J Pathol 144, 337–47 (1994).
Lindahl B, Eriksson L, Lindahl U, Biochem J 306, 177–84 (1995).
Thomas SS, Plenkiewicz J, Ison ER, Bols M, Zou W, Szarek WA, Kisilevsky R, Biochim Biophys Acta 1272, 37–48 (1995).
Dow KE, Riopelle RJ, Szarek WA, Bols M, Ison ER, Plenkiewicz J, Lyon A, Kisilevsky R, Biochim Biophys Acta 1156, 7–14 (1992).
Berkin A, Szarek WA, Kisilevsky R, Carbohydr Res 326, 250–63 (2000).
Ledbetter SR, Hassell JR, Arch Biochem Biophys 246, 403–10 (1986).
Fritz TA, Lugemwa FN, Sarkar AK, Esko JD, J Biol Chem 269, 300–7 (1994).
Lugemwa FN, Esko JD, J Biol Chem 266, 6674–7 (1991).
Lugemwa FN, Esko JD, Carbohydr Res 239, 285–90 (1993).
Miao HQ, Fritz TA, Esko JD, Zimmermann J, Yayon A, Vlodavsky I, J Cell Biochem 57, 173–84 (1995).
Parry G, Farson D, Cullen B, Bissell MJ, In Vitro 24, 1217–22 (1988).
Sudhakaran R, Sinn W, von Figura K, Hoppe-Seyler's Z Physiol Chem 362, 39–46 (1981).
Margolis RK, Goossen B, Tekotte H, Hilgenberg L, Margolis RU, J Cell Sci 99, 237–46 (1991).
van Kuppevelt THSM, Van de Lest CHA, Versteeg EMM, Dekhuijzen PNR, Veerkamp JH, Amer J Respir Cell Mol Biol 16, 75–84 (1997).
Bernacki RJ, Korytnyk W, Development of membrane sugar and nucleotide sugar analogs as potential inhibitors or modifiers of cellular glycoconjugates. In The glycoconjugates, Vol. 4, Part B, edited by Horowitz MI (Academic Press, New York, 1982), pp. 245–63.
Sharma M, Bernacki RJ, Paul B, Korytnyk W, Carbohydr Res 198, 205–21 (1990).
Paul B, Bernacki RJ, Korytnyk W, Carbohydr Res 80, 99–115 (1980).
Subrahmanyan L, Kisilevsky R, Scand J Immunol 27, 251–60 (1988).
Kisilevsky R, Subrahmanyan L, Lab Invest 66, 778–85 (1992).
Hronowski L, Anastassiades TP, J Biol Chem 255, 9210–17 (1980).
Inouye Y, Onodera K, Kitaoka S, Ochiai H, J Am Chem Soc 79, 4218–22 (1957).
Hudson CS, Dale JK, J Am Chem Soc 38, 1431–36 (1916).
Berkin A, Szarek MA, Plenkiewicz J, Szarek WA, Kisilevsky R, Carbohydr Res 325, 30–45 (2000).
Horton D, Wolfrom ML, J Org Chem 27, 1794–1800 (1962).
Thomas SS, M.Sc. Thesis, Queen's University, Kingston, Ontario (1991).
Bernacki RJ, Sharma M, Porter NK, Rustum Y, Paul B, Korytnyk W, Supramol Struct 7, 235–50 (1977).
Decker K, Keppler D, In Progress in liver disease, Vol. 4, edited by Popper H, Schaffner F (Grune & Stratton, New York, 1972), pp. 183–99.
Decker K, Keppler D, Rev Physiol Biochem Pharmacol 71, 77–106 (1974).
Roeser KR, Legler G, Biochim Biophys Acta 657, 321–33 (1981).
McCarter JD, Adam MJ, Withers SG, Biochem J 286, 721–7 (1992).
Berkin A, Szarek WA, Kisilevsky R, Carbohydr Res 337, 37–44 (2002).
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Berkin, A., Szarek, W.A. & Kisilevsky, R. Biological evaluation of a series of 2-acetamido-2-deoxy-D-glucose analogs towards cellular glycosaminoglycan and protein synthesis in vitro . Glycoconj J 22, 443–451 (2005). https://doi.org/10.1007/s10719-005-5060-1
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DOI: https://doi.org/10.1007/s10719-005-5060-1