ABSTRACT
Teredinibacter turnerae is a cultivable cellulolytic Gammaproeteobacterium (Cellvibrionaceae) that commonly occurs as an intracellular endosymbiont in the gills of wood-eating bivalves of the family Teredinidae (shipworms). The genome of T. turnerae encodes a broad range of enzymes that deconstruct cellulose, hemicellulose, and pectin and contribute to lignocellulose digestion in the shipworm gut. However, the mechanism by which symbiont-made enzymes are secreted by T. turnerae and subsequently transported to the site of lignocellulose digestion in the shipworm gut is incompletely understood. Here, we show that T. turnerae cultures grown on carboxymethyl cellulose (CMC) produce outer membrane vesicles (OMVs) that contain a variety of proteins identified by LC-MS/MS as carbohydrate-active enzymes with predicted activities against cellulose, hemicellulose, and pectin. Reducing sugar assays and zymography confirm that these OMVs retain cellulolytic activity, as evidenced by hydrolysis of CMC. Additionally, these OMVs were enriched with TonB-dependent receptors, which are essential to carbohydrate and iron acquisition by free-living bacteria. These observations suggest potential roles for OMVs in lignocellulose utilization by T. turnerae in the free-living state, in enzyme transport and host interaction during symbiotic association, and in commercial applications such as lignocellulosic biomass conversion.
ABSTRACT
Shipworms are marine wood-boring bivalve mollusks (family Teredinidae) that harbor a community of closely related Gammaproteobacteria as intracellular endosymbionts in their gills. These symbionts have been proposed to assist the shipworm host in cellulose digestion and have been shown to play a role in nitrogen fixation. The genome of one strain of Teredinibacter turnerae, the first shipworm symbiont to be cultivated, was sequenced, revealing potential as a rich source of polyketides and nonribosomal peptides. Bioassay-guided fractionation led to the isolation and identification of two macrodioloide polyketides belonging to the tartrolon class. Both compounds were found to possess antibacterial properties, and the major compound was found to inhibit other shipworm symbiont strains and various pathogenic bacteria. The gene cluster responsible for the synthesis of these compounds was identified and characterized, and the ketosynthase domains were analyzed phylogenetically. Reverse-transcription PCR in addition to liquid chromatography and high-resolution mass spectrometry and tandem mass spectrometry revealed the transcription of these genes and the presence of the compounds in the shipworm, suggesting that the gene cluster is expressed in vivo and that the compounds may fulfill a specific function for the shipworm host. This study reports tartrolon polyketides from a shipworm symbiont and unveils the biosynthetic gene cluster of a member of this class of compounds, which might reveal the mechanism by which these bioactive metabolites are biosynthesized.
