Born 1928, Jerusalem; Ph.D. 1956, Hebrew Univ.; Lect. 1961; Sen. Lect. 1965; Assoc. Prof. 1971; Prof. 1977; Emeritus October 1996.
Biochemistry and molecular biology of cellulose biosynthesis.
Biochemistry and molecular biology of cellulose biosynthesis. Funding: Weyerhaeuser Co. (USA); Israel Ministry of Science and Arts.
Abstracts of Current Research:
Biochemistry and molecular biology of cellulose biosynthesis:
The major focus of investigation is the biochemistry and molecular biology of cellulose (?-1,4-glucan) biogenesis. This carbohydrate merits special consideration in the current global concern for the environment, as it is the world's most abundant natural, renewable and biodegradable polymer. The gram negative bacterium Acetobacter xylinum has proved to be an effective model system, as it produces cellulose structurally similiar to that produced by higher plants, but in a much purer form as an extracellular deposit. We have found that in this organism, cellulose biogenesis is governed by a multicomponent regulatory system, which was elucidated following isolation of a catalytically active cellulose synthase (CS) and the discovery of the novel dinucleotide cyclic diguanylic acid (c-di-GMP) as its allosteric effector. Our results at the biochemical level directly enabled molecular cloning of genes involved in cellulose synthesis, including those responsible for formation of the substrate UDP-glucose, formation and degradation of c-di-GMP and those encoding the essential genes of the CS itself. Molecular manipulations of cloned genes and insertion into mutant strains with known deficiencies are being conducted to determine exact gene functions, with the goal of optimizing cellulose production. An additional protein component of the system which appears to regulate the intracellular concentration of c-di-GMP has been discovered and cloning of its corresponding gene is being pursued.
In contrast to the bacterial system, in vitro activity of a plant CS has yet to be convincingly demonstrated, although a large body of evidence has been accumulated indicating homologous elements at both the protein and nucleic acid levels. Discovery of the plant CS will likely have immediate impact in the field of biotechnology for crop improvement. Interestingly, efforts to produce cellulose from plant extracts yield the related polymer callose (?-1,3-glucan) which is produced by plants under conditions of stress and in specialized structures such as pollen tubes. We have discovered an allosteric activator of the callose synthase which has been characterized as ?-furfuryl-?-glucoside, and its mode of action has also been elucidated.
The unique compound c-di-GMP may provide the missing link between the plant and bacterial systems with regard to cellulose formation. Diguanylate cyclase, the enzyme which catalyzes formation of this nucleotide, has been found to be inhibited by a unique saponin which was first isolated from a plant source and subsequently found in A. xylinum. We are currently searching for the diguanylate cyclase enzyme itself and its related genes in plant systems.
In a parallel approach to identify a plant CS gene, the cotton fiber is being used as a model system, as this plant produces vast amounts of cellulose in a tissue specific and developmentally related manner. A subtractive technique has been developed to identify cDNA clones which are expressed exclusively in tissues actively engaged in cellulose synthesis.
Wong, H.C., Fear, A.L., Calhoon, R.D., Eichinger, G.H., Mayer, R., Amikam, D. Benziman, M., Gelfand, D.H., Meade, J.H., Emerick, A.W., Bruner, R., Ben-Bassat, A. and Tal, R. (1990) Genetic organization of the cellulose synthase operon in Acetobacter xylinum. Proc. Natl. Acad. Sci. USA, 87: 8130-8134.
Mayer, R., Ross, P., Weinhouse, H., Amikam, D., Volman, G., Ohana, P., Calhoon, R.D., Wong, H.C., Emerick, A.W. and Benziman, M. (1991) Polypeptide composition of bacterial cyclic diguanylic acid-dependent cellulose synthase and the occurrence of immunologically crossreacting proteins in higher plants. Proc. Natl. Acad. Sci USA, 88: 5472-5476.
Ross, P., Mayer, R. and Benziman, M. (1991) Cellulose biosynthesis and function in bacteria. Microbiological Reviews, 55: 35-58.
Amor, Y., Mayer, R., Delmer, D.P. and Benziman, M. (1991) Evidence for a c-di-GMP dependent cellulose synthase in plants. The Plant Cell, 3: 989-995.
Ohana, P., Delmer, D.P., Steffens, J.C., Matthews, D.E., Mayer, R. and Benziman, M. (1991) ?-furfuryl-?-glucoside. An endogenous activator of higher plant UDP-glucose: (1->3)-?-glucan synthase. J. Biol. Chem., 266: 13742-13745.
Ohana, P., Delmer, D.P., Volman, G., Steffens, J.C., Matthews D.E. and Benziman, M. (1991) ?-furfuryl-?-glucoside: An endogenous activator of higher plant UDP-glucose:(1->3)-?-glucan synthase. Biological activity, distribution and in vitro synthesis. Plant Physiol., 98: 708-715.
Ohana, P., Benziman, M. and Delmer, D.P. (1993) Stimulation of callose synthesis in vivo correlates with changes in intracellular distribution of the callose synthase activator ?-furfuryl-?-glucoside. Plant Physiol., 101: 187-191.
Volman, G., Ohana, P. and Benziman, M. (1995) Biochemistry and molecular biology of cellulose biosynthesis. Carbohydrates in Europe, 12: 20-27.
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