Effect of over-expression of phasin gene from Aeromonas hydrophila on biosynthesis of copolyesters of 3-hydroxybutyrate and 3-hydroxyhexanoate

The gene phaP Ah, encoding the protein phasin that is associated with poly(3-hydroxybutyrate- co-3-hydroxyhexanoate) (PHBHHx) granule of Aeromonas hydrophila 4AK4, was cloned and characterized. Recombinant strains harboring additional copies of the phasin gene ( phaP Ah) and the polyhydroxyalkanoate (PHA) synthase gene ( phaC Ah) accumulated PHBHHx copolyesters consisting of 21 mol% 3-hydroxyhexanoate (3HHx) as compared to 14 mol% 3HHx produced by wild type strain. The molecular weight of PHBHHx produced by the above recombinants was lower than that obtained from the wild type strain grown under similar conditions. Over-expression of phaP Ah led to the production of more PHA granules but with reduced sizes. SDS–PAGE showed that PhaP Ah was the predominant protein present in the PHBHHx granules. The RT-PCR results suggested that phasin PhaP Ah, regulated phaC Ah gene at the transcription level. Gene PhaP We from Wautersia eutropha (formerly Ralstonia eutropha; encoding a 20 kDa protein with low amino acid homology to the A. hydrophila 13 kDa protein) cloned into A. hydrophila 4AK4 exhibited similar effects on PHBHHx production and PHBHHx composition. These data suggest that the phasins could represent a protein family possessing similar functions but different structures.