Gene cloning and enzyme structure modeling of the Aspergillus oryzae N74 fructosyltransferase

Abstract

The fructooligosaccharides (FOS) represent an important source of prebiotic compounds that are widely used as an ingredient in functional foods. Recently, the strain Aspergillus oryzae N74 was reported as a potential microorganism for the industrial production of FOS, due to its high yields of FOS production. In this work, we used a PCR-cloning strategy to clone the A. oryzae N74 ftase gene as a previous step for recombinant enzyme production. Ftase showed a 1630 bp size with a 99% similarity with other A. oryzae strains and between 1 to 68% identities with other Aspergillus strains. This gene encodes for a 525 amino acids protein with 99% similarity with other A. oryzae strains and between 11 to 69% similarities with other Aspergillus strains. Finally, an A. oryzae N74 FTase tertiary structure model was predicted base on its similarity with other glycoside hydrolase 32 family members. The active site was located inside the β-propeller domain and was formed for non-charged polar and charged amino acids. In summary, these results shows the high level of sequence conservation between A. oryzae strains and represent a first step towards the development of a FOS production industrial process using recombinant microorganism carrying the ftase gene from A. oryzae N74.