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Structural and functional features of fructansucrases present in Leuconostoc mesenteroides ATCC 8293

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Abstract

Glycosyltransferases produced by Leuconostoc mesenteroides subsp. mesenteroides ATCC 8293 (equivalent to NRRL B-1118) were identified. Two glucansucrases and one fructansucrases were observed in batch culture while levC and levL genes, corresponding to two fructansucrases, were isolated from information obtained from the released draft sequence of this Leuconostoc strain genome and cloned in Escherichia coli. The recombinant enzymes were shown to be fructansucrases producing a polymer identified by NMR as levan, confirming our recent report stating that these are also mosaic levansucrases bearing structural features of glucansucrases in the amino and carboxy terminal regions, as is also the case of inulosucrase (IslA) from Leuconostoc citreum CW28 and levansucrase (LevS) from L. mesenteroides NRRL B-512F. The recombinant levansucrase LevC was purified and characterized in terms of pH, temperature, and kinetic properties. The enzyme exhibits Michaelis–Menten kinetic properties with a K m = 27.3 mM and a k cat = 282.9 s−1. This levansucrase behaves mainly as a transferase as only 30% of the substrate is hydrolyzed in a wide range of sucrose concentrations, with higher hydrolytic activities at low substrate concentrations. With this report we experimentally confirm the unusual structural pattern displayed by fructansucrases present in Leuconostoc species that group as a novel sub family of fructansucrases.

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Acknowledgments

This project was supported by Consejo Nacional de Ciencia y Tecnología (CONACyT) no. 40609-Z and by UNAM (PAPIIT No. IN228006–3). We thank Fernando González, Maria Elena Rodriguez and Ricardo Ciria for technical assistance. We also thank Eugenio López, Paul Gaytán, and Jorge Yáñez, for the primer synthesis and sequences analysis

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  1. Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Avenida Universidad 2001 Col. Chamilpa, Cuernavaca, Morelos, 62250, Mexico

    Clarita Olvera, Sara Centeno-Leija & Agustín López-Munguía

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  1. Clarita Olvera
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  2. Sara Centeno-Leija
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  3. Agustín López-Munguía
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Correspondence to Agustín López-Munguía.

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Olvera, C., Centeno-Leija, S. & López-Munguía, A. Structural and functional features of fructansucrases present in Leuconostoc mesenteroides ATCC 8293. Antonie van Leeuwenhoek 92, 11–20 (2007). https://doi.org/10.1007/s10482-006-9128-0

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  • DOI: https://doi.org/10.1007/s10482-006-9128-0

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