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Synthesis of chitin by particulate preparations from Aspergillus flavus

  • Biochemistry
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Abstract

Cell-free extracts from Aspergillus flavus catalyzed the synthesis of chitin from UDP-GlcNAc. Most of the activity was associated with membrane-rich fractions whereas no activity was detected in the cell walls. Chitin synthetase was activated by fungal acid proteases; animal and plant proteases destroyed it. Upon incubation at 0 C and 28 C chitin synthetase was inactivated, probably by the action of proteases present in the particulate preparations. Maximal activity was obtained at pH 6.6–7.1 and 15 C. Arrhenius plot showed a biphasic curve with the transition at 7 C. E values were 3300 Kcal/mole above this temperature and 15500 Kcal/mole below it. The enzyme was activated by GlcNAc and required a divalent metal, the most active being Mg++. By plotting v vs UDP-GlcNAc concentration a sigmoidal curve was obtained. Km calculated at high substrate concentrations was 20mm. Chitin synthetase was competitively inhibited by polyoxin D (Ki 6.5 μm) and UDP (Ki 1.35mm), the latter giving complex kinetics.

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References

  • Bartnicki-García, S. and Nickerson, W. J. 1962. Nutrition, growth, and morphogenesis of M. rouxii. — J. Bacteriol. 84: 841–858.

    Google Scholar 

  • Bartnicki-García, S. and Lippman, E. 1972. Inhibition of Mucor rouxii by polyoxin D: effects on chitin synthetase and morphological development. — J. Gen. Microbiol. 71: 301–309.

    Google Scholar 

  • Berger, L. R. and Reynolds, D. M. 1958. The chitinase system of a strain of Streptomyces griseus. — Biochim. Biophys. Acta 29: 522–534.

    Google Scholar 

  • Cabib, E. and Farkas, V. 1971. The control of morphogenesis: an enzymatic mechanism for the initiation of septum formation in yeast. — Proc. Nat. Acad. Sci. U.S.A. 68: 2052–2056.

    Google Scholar 

  • Camargo, E. P., Dietrich, C. P., Sonneborn, D. and Strominger, J. L. 1967. Biosynthesis of chitin in spores and growing cells of Blastocladiella emersonii. — J. Biol. Chem. 242: 3121–3128.

    Google Scholar 

  • Carey, F. G. 1965. Chitin synthesis in vitro by crustacean enzymes. — Comp. Biochem. Physiol. 16: 155–158.

    Google Scholar 

  • Dixon, M. 1953. The determination of enzyme inhibitor constants. — Biochem. J. 55: 170–171.

    Google Scholar 

  • Dixon, M. and Webb, E. C. 1964. Enzymes, 2nd Ed., p. 158. — Longman Group Limited, London.

    Google Scholar 

  • Endo, A., Kakiki, K. and Misato, T. 1970. Mechanism of action of the antifungal agent polyoxin D. — J. Bacteriol. 104: 189–196.

    Google Scholar 

  • Endo, A. and Misato, T. 1969. Polyoxin D, a competitive inhibitor of UDP-N-acetylglucosamine: chitin N-acetylglucosaminyl transferase in Neurospora crassa. — Biochem. Biophys. Res. Commun. 37: 718–722.

    Google Scholar 

  • Glaser, L. and Brown, D. H. 1957. The synthesis of chitin in cell-free extracts of Neurospora crassa. — J. Biol. Chem. 228: 729–742.

    Google Scholar 

  • Guerrero-Caballero, R. and Ruiz-Herrera, J. 1971. A kinetic analysis of the citrate permeability system from Aerobacter aerogenes. — Rev. Lat. Amer. Microbiol. 13: 297–302.

    Google Scholar 

  • Hori, M., Kakiki, K. and Misato, T. 1974a. Further study of the relation of polyoxin structure to chitin synthetase inhibition. — Agr. Biol. Chem. 38: 691–698.

    Google Scholar 

  • Hori, M., Kakiki, K. and Misato, T. 1974b. Interaction between polyoxin and active center of chitin synthetase. — Agr. Biol. Chem. 38: 699–705.

    Google Scholar 

  • Isono, K., Asahi, K. and Suzuki, S. 1969. Studies on polyoxins, antifungal antibiotics. XIII. The structure of polyoxins. — J. Am. Chem. Soc. 91: 7490–7505.

    Google Scholar 

  • Jan, Y. N. 1974. Properties and cellular localization of chitin synthetase in Phycomyces blakesleeanus. — J. Biol. Chem. 249: 1973–1979.

    Google Scholar 

  • Jaworski, E. G., Wang, L. C. and Carpenter, W. D. 1965. Biosynthesis of chitin in cell-free extracts of Venturia inaequalis. — Phytopathology 55: 1309–1312.

    Google Scholar 

  • Jeuniaux, C. 1966. Chitinases, p. 644–650. In Methods in Enzymology, Vol. VIII. — Academic Press Inc., New York and London.

    Google Scholar 

  • Keller, F. A. and Cabib, E. 1971. Chitin and yeast budding. Properties of chitin synthetase from Saccharomyces carlsbergensis. — J. Biol. Chem. 246: 160–166.

    Google Scholar 

  • Larson, A. D. and Kallio, R. E. 1954. Purification and properties of bacterial urease. —J. Bacteriol. 68: 67–73.

    Google Scholar 

  • Levy, H. M., Sharon, N. and Koshland, D. E. Jr. 1959a. A mechanism for the effects of dinitrophenol and temperature on the hydrolytic activity of myosin. — Biochim. Biophys. Acta 33: 288–289.

    Google Scholar 

  • Levy, H. M., Sharon, N. and Koshland, D. E. Jr. 1959b. Purified muscle proteins and the walking rate of ants. — Proc. Nat. Acad. Sci. U.S.A. 45: 785–791.

    Google Scholar 

  • Lowry, O. H., Rosebrough, N. J., Farr, A. L. and Randall, R. J. 1951. Protein measurement with the Folin phenol reagent. — J. Biol. Chem. 193: 265–275.

    Google Scholar 

  • Magaña-Plaza, I., Montes, C. and Ruiz-Herrera, J. 1971. Purification and biochemical characteristics of urease from Proteus rettgeri. — Biochim. Biophys. Acta 242: 230–237.

    Google Scholar 

  • Massey, V. 1953. Studies on fumarase. 3. The effect of temperature. — Biochem. J. 53: 72–79.

    Google Scholar 

  • McMurrough, I. and Bartnicki-García, S. 1971. Properties of a particulate chitin synthetase from Mucor rouxii. — J. Biol. Chem. 246: 4008–4016.

    Google Scholar 

  • McMurrough, I., Flores-Carreón, A. and Bartnicki-García, S. 1971. Pathway of chitin synthesis and cellular localization of chitin synthetase in Mucor rouxii. — J. Biol. Chem. 246: 3999–4007.

    Google Scholar 

  • Ohta, N., Kakiki, K. and Misato, T. 1970. Studies on the mode of action of polyoxin D. Part II. Effect of polyoxin D on the synthesis of fungal cell wall chitin. — Agr. Biol. Chem. 34: 1224–1234.

    Google Scholar 

  • Overath, P., Schairer, H. U., Hill, F. F. and Hirsch, I. L. 1971. Structure and function of hydrocarbon chains in bacterial phospholipids, p. 161. In D. F. Hölzl Wallach and H. Fisher, (eds.), The dynamic structure of cell membranes. — Springer-Verlag, New York.

    Google Scholar 

  • Peberdy, J. F. and Moore, P. M. 1975. Chitin synthase in Mortierella vinacea: properties, cellular location and synthesis in growing cultures. — J. Gen. Microbiol. 90: 228–236.

    Google Scholar 

  • Porter, C. A. and Jaworski, E. G. 1966. The synthesis of chitin by particulate preparations of Allomyces macrogynus. — Biochemistry 5: 1149–1154.

    Google Scholar 

  • Roseman, S. and Ludowieg, J. 1954. N-acetylation of the hexosamines. — J. Am. Chem. Soc. 76: 301–302.

    Google Scholar 

  • de Rousset-Hall, A. and Gooday, G. W. 1975. A kinetic study of a solubilized chitin synthetase preparation from Coprinus cinereus. — J. Gen. Microbiol. 89: 146–154.

    Google Scholar 

  • Ruiz-Herrera, J. 1967. Chemical components of the cell wall of Aspergillus species. —Arch. Biochem. Biophys. 122: 118–125.

    Google Scholar 

  • Ruiz-Herrera, J. and Bartnicki-García, S. 1974. Synthesis of cell wall microfibrils in vitro by a “soluble” chitin synthetase from Mucor rouxii. — Science 186: 357–359.

    Google Scholar 

  • Ruiz-Herrera, J. and Bartnicki-García, S. 1975. Biochemistry of dimorphism: proteolytic activation and destruction of chitin synthetase from Mucor rouxii. — Abstr. Ann. Meet. Amer. Soc. Microbiol., 130.

  • Ruiz-Herrera, J., Sing, V. O., van der Woude, W. J. and Bartnicki-García, S. 1975. Microfibril assembly by granules of chitin synthetase. — Proc. Nat. Acad. Sci. U.S.A. 72: 2706–2710.

    Google Scholar 

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Author notes

  1. E. López-Romero (Predoctoral Fellow)

    Present address: Consejo Nacional de Ciencia y Tecnologia, Mexico, México

  2. J. Ruiz-Herrera (Professorial Fellow)

    Present address: Comisión de Operación y Fomento de las Actividades Académicas of the I.P.N., México, México

Authors and Affiliations

  1. Departamento de Microbiología, Escuela Nacional de Ciencias Biológicas, I.P.N., 17, México, D.F., México

    E. López-Romero (Predoctoral Fellow) & J. Ruiz-Herrera (Professorial Fellow)

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  1. E. López-Romero
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  2. J. Ruiz-Herrera
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Additional information

This work was supported by grants No. 020 and 847 of the Consejo Nacional de Ciencia y Tecnología, México.

Part of this work was carried out while the authors were research visitors at the Department of Plant Pathology, University of California, Riverside.

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López-Romero, E., Ruiz-Herrera, J. Synthesis of chitin by particulate preparations from Aspergillus flavus . Antonie van Leeuwenhoek 42, 261–276 (1976). https://doi.org/10.1007/BF00394123

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