Abstract

Fruit ripening is a coordinated event and involves several changes promoting the development of an attractive fruit to eat. Particularly, texture modifications leading to fruit softening are strongly related to cell wall metabolism, and several enzymes acting on cell wall polysaccharides have been identified. Fragaria chiloensis (L.) Mill. is a Chilean endemic fruit with excellent organoleptic properties, however its fast softening during ripening is responsible of its brief postharvest shelf-life. Previous research has concluded that the main cell wall polysaccharide lost during ripening are the pectins, primarily rhamnogalacturonan I (RG-I). Until now there is information about enzymes acting on RG-I side chains, nevertheless little is known about enzymes acting on RG-I backbone. From a transcriptome of F. chiloensis fruit a contig encoding to a rhamnogalacturonan endolyase (RGL4, Polysaccharide lyase 4 family; EC number 4.2.2.23) was identified whose FPKM values increased during its ripening. Through the catalytic β-elimination mechanism RGL4 breaks the α-1,4-glycosidic bond between rhamnose and galacturonic acid of RG-I backbone. The predicted RGL4 sequence consists of 676 amino acid residues, including a signal peptide. Two putative N-glycosylation sites are predicted. It shares 24% identity with RGL4 from the saprophytic fungus Aspergillus aculeatus, and also the three characteristic domains in the tertiary structure are conserved as well as the key catalytic residues (K217 and H276). Understanding the role played by this enzyme in cell wall disassembly will contribute to a comprehensive knowledge of the molecular mechanisms involved in the ripening of F. chiloensis fruit.