Identification and functional characterization of a fructose-inducible phosphotransferase system in Azospirillum brasilense Sp7

Abstract

Plant growth-promoting rhizobacterium Azospirillum brasilense Sp7 utilizes fructose efficiently via a fructose phosphotransferase system (Fru-PTS). Its genome encodes two putative Fru-PTS, each consisting of FruB (EIIA), FruK (Pfk), and FruA (EIIBC) proteins. We compared the proteomes of A. brasilense Sp7 grown with malate or fructose as sole carbon source, and noticed upregulation of the constituent proteins of Fru-PTS1 only on fructose. Inactivation of fruA gene of both the Fru-PTS showed that Fru-PTS1 is the main PTS involved in fructose utilization. Overexpression of fruA1 in A. brasilense Sp7 enhanced its growth on fructose showing improved consumption of fructose. This suggested that fructose utilization in A. brasilense Sp7 is limited due to the limitation of EIIBC component. A FruR-type regulator, encoded divergently to the Fru-PTS1 operon, was required for chemotaxis toward fructose. Although not an absolute necessity for the growth of fructose, FruR was required for the optimal growth of fructose. The fruB1 promoter was activated by fructose and repressed by malate, but FruR does not seem to regulate its expression. A 27-nucleotide stem-loop structure located between the −125 and −99 promoter proximal region of fruB1 was involved in fructose inducibility and malate repression. Fructose also upregulated several proteins involved in the biogenesis of a Type 6 secretion system. Here, we have shown that A. brasilense Sp7 was able to inhibit the growth of Escherichia coli and Agrobacterium tumefaciens in the presence of fructose, and that an intact T6SS was required for contact-dependent growth inhibition of the two Gram-negative bacteria.