Galactofuranosidase from JHA 19 Streptomyces sp:. subcloning and biochemical characterization

Mateja Senicar (1, 2) , Laurent Legentil (3) , Vincent Ferrières (3) , Eliseeva V. Svetlana (1) , Stéphane Petoud (1) , Kaoru Takegawa (4) , Pierre Lafite (2) , Richard Daniellou (2)

1 CBM - Centre de biophysique moléculaire
2 ICOA - Institut de Chimie Organique et Analytique
3 ISCR - Institut des Sciences Chimiques de Rennes
4 Kyushu University

Despite the crucial role of the rare galactofuranose (Galf) in many pathogenic micro-organisms and our increased knowledge of its metabolism, there is still a lack of recombinant and efficient galactofuranoside hydrolase available for chemo-enzymatic synthetic purposes of specific galactofuranosyl-conjugates. Subcloning of the Galf-ase from JHA 19 Streptomyces sp. and its further overexpression lead us to the production of this enzyme with a yield of 0.5 mg/L of culture. It exhibits substrate specificity exclusively towards pNP beta-D-Galf, giving a K-M value of 250 mu M, and the highest enzymatic efficiency ever observed of 14 mM(-1) s(-1). It proved to be stable to temperature up to 60 degrees C and to at least 4 freeze-thaw's cycles. Thus, Galf-ase demonstrated to be an efficient and stable biocatalyst with greatly improved specificity toward the galactofuranosyl entity, thus paving the way to the further development of transglycosylation and thioligation reactions.

Keywords: beta-D-galactofuranosidase; Subcloning; Glycoside hydrolase; Thio-beta-D-galactofuranoside inhibition

Domains: Life Sciences [q-bio]; Biotechnology

Mateja Senicar, Laurent Legentil, Vincent Ferrières, Eliseeva V. Svetlana, Stéphane Petoud, et al.. Galactofuranosidase from JHA 19 Streptomyces sp:. subcloning and biochemical characterization. Carbohydrate Research, 2019, 480, pp.35-41. ⟨10.1016/j.carres.2019.05.011⟩. ⟨hal-02177806⟩