Identification of an Extracellular Poly(3-hydroxybutyrate) Depolymerase in the Genus Alteromonas and Its Phylogenetic...

Poly(3-hydroxybutyrate), P(3HB), is an aliphatic polyester that is susceptible to biodegradation even in marine environments. The high biodegradability of P(3HB) can be attributed to the presence in the environment of extracellular P(3HB) depolymerase (PhaZ), the initial enzyme involved in P(3HB) degradation. In this study, we aimed to identify the gene encoding PhaZ in the marine P(3HB)-degrading bacterium Alteromonas sp. D210916BOD_24, which was previously isolated. First, we conducted genome analysis of the strain, revealing that the strain possesses a PhaZ homolog. It possesses a catalytic domain with a lipase box near the center, a substrate-binding domain comprising two regions, and a fibronectin type III linker domain, which fit the marine bacterial domain pattern. Disruption of the PhaZ homolog gene caused the strain to lose its P(3HB) degradation ability. The recombinant PhaZ homolog protein exhibited significant activity toward P(3HB). The results indicated that the PhaZ homolog indeed functions as PhaZ in Alteromonas sp. D210916BOD_24. Furthermore, we focused on the distribution and genomic placement of PhaZ homolog genes. The PhaZ homolog was present in 4/20 Alteromonas strains examined, with Alteromonas sp. D210916BOD_24 showing substantial divergence from the other three strains in the 16S rDNA-based phylogenetic tree. The gene arrangement around the PhaZ homolog gene was clearly different between Alteromonas sp. D210916BOD_24 and the others. Additionally, large-scale gene locations and orientations exhibited considerable differences. These findings suggest that the horizontal transfer of PhaZ homolog genes occurred after a certain degree of species division.