Marine algae including red algae (Rhodophyta) have recently been considered as a promising resource complementing land plants for bioenergy production. The major composite of red algal cell wall is an agar polysaccharide. Biological process degrades the agar polysaccharide into monomeric sugars by several steps, which can be a platform for designing biochemical red algal biomass deconstruction process. Various agarases are involved in each step of the decomposition. We collected all known agarase genes from public database and reclassified them using a domain-based network. From our model, we identified candidate agarases from all known sequenced genome. Among selected genes, several were expressed in E. coli to test their predicted activity. Among them, two agarases belonging to glycohydrolase family 50 (GH50) from Saccharophagus degradans were overexpressed as soluble. We deduced active sites from the sequence and structure analysis. The enzymes were identified as exo-type agarase producing neoagarobiose as a reaction product. Although two enzymes show high sequence similarity, the level of activity was much different. We suggest that various agarases collected from our approach can be used in biochemical deconstruction process of red algae biomass.