The levansucrase puzzle: how to combine structure and biochemistry to understand the biological function

Ivan Polsinelli1, Rosanna Caliandro1, Marco Salomone-Stagni1, Nicola Demitri2, Martin Rejzek3, Robert A. Field3, Stefano Benini1,*

1 Bioorganic Chemistry and Bio-Crystallography laboratory (B2Cl), Faculty of Science and Technology, Free University of Bolzano, Piazza Università 5, 39100 Bolzano, Italy

2 Elettra-Sincrotrone Trieste, S.S. 14 Km 163.5 in Area Science Park, Basovizza, Trieste 34149, Italy

3 Department of Biological Chemistry, John Innes Centre, Norwich Research Park, Norwich, NR47UH, United Kingdom

* corresponding author: stefano.benini@unibz.it

The Gram-negative bacterium Erwinia amylovora is the etiological agent of fire blight, a devastating disease which affects Rosaceae. Activity of levansucrase has been correlated with the virulence of E. amylovora. E. amylovora levansucrase defective mutants, showed retarded development of necrotic symptoms on pear seedlings inoculated with the mutated strains [1]. The plant pathogen E. amylovora is related to the epiphytic bacterium Erwinia tasmaniensis [2, 3].

To understand the role of this enzyme in the two bacteria, the levansucrase from E. tasmaniensis (EtLsc) has been compared with the homologous enzyme from E. amylovora (EaLsc) [4]. The enzymatic activity was characterized by High Performance Anion Exchange Chromatography coupled with Pulsed Amperometric Detector (HPAEC-PAD). The comparison of the products profile highlights an increased efficiency of EtLsc than EaLsc. The crystal structure of EtLsc reveals structural peculiarities causing the different product profiles of the two homologues.

Based on our results, we propose that the role of this enzyme in the life cycle of the two bacteria is most likely related to survival, rather than linked to the virulence of the plant pathogen E. amylovora [5].

1. Geier, G., Geider, K., 1993. Characterization and influence on virulence of the levansucrase gene from the fireblight pathogen Erwinia-amylovora. Physiological and Molecular Plant Pathology 42, pp. 387-404.

2. Geider, K., Auling, G., Du, Z., Jakovljevic, V., Jock, S., Volksch, B., 2006. Erwinia tasmaniensis sp. nov., a non-phytopathogenic bacterium from apple and pear trees. Int. J. Syst. Evol. Microbiol. 56, pp. 2937-2943.

3. Borruso, L., Salomone-Stagni, M., Polsinelli, I., Schmitt, A.O., Benini, S., 2017. Conservation of Erwinia amylovora pathogenicity-relevant genes among Erwinia genomes. Arch. Microbiol. 199, pp. 1335-1344.

4. Caputi, L., Nepogodiev, S.A., Malnoy, M., Rejzek, M., Field, R.A., Benini, S., 2013. Biomolecular characterization of the levansucrase of Erwinia amylovora, a promising biocatalyst for the synthesis of fructooligosaccharides. J. Agric. Food Chem. 61, pp. 12265-12273.

5. Polsinelli, I., Caliandro, R., Salomone-Stagni, M., Demitri, N., Rejzek, M., Field, R.A., Benini, S., 2019. Comparison of the Levansucrase from the epiphyte Erwinia tasmaniensis vs its homologue from the phytopathogen Erwinia amylovora. Int. J. Biol. Macromol. 127, pp. 496-501.