Comparison of four phaC genes from Haloferax mediterranei and their function in different PHBV copolymer biosyntheses in Haloarcula hispanica

With the aid of the genome sequence of Hfx. mediterranei CGMCC 1.2087, three additional phaC genes (designated phaC1, phaC2, and phaC3) were identified, which encoded putative PhaCs. Like PhaCHme (54.8 kDa), PhaC1 (49.7 kDa) and PhaC3 (62.5 kDa) possessed the conserved motifs of type III PHA synthase, which was not observed in PhaC2 (40.4 kDa). Furthermore, the longer C terminus found in the other three PhaCs was also absent in PhaC2. Reverse transcription PCR (RT-PCR) revealed that, among the four genes, only phaCHme was transcribed under PHA-accumulating conditions in the wild-type strain. However, heterologous coexpression of phaEHme with each phaC gene in Haloarcula hispanica PHB-1 showed that all PhaCs, except PhaC2, could lead to PHBV accumulation with various 3HV fractions. The three kinds of copolymers were characterized using gel-permeation chromatography (GPC), differential scanning calorimetry (DSC), and thermogravimetric analysis (TGA). Their thermal properties changed with the variations in monomer composition as well as the different molecular weights (Mw), thus might meet various application requirements.We discover three cryptic phaC genes in Hfx. mediterranei, and demonstrate that genetic engineering of these newly identified phaC genes has biotechnological potential for PHBV production with tailor-made material properties.Polyhydroxyalkanoates (PHAs) are polymers of hydroxyalkanoates and are synthesized by various bacteria and some haloarchaeal strains [1]. The molecular weight of PHAs is generally distributed in the range of 50,000-1,000,000 Da, which is sufficient to provide these polymers with similar characteristics as conventional plastics [2]. However, due to the high production cost of PHAs, semi-commercial production has been realized only for the two common polyesters, poly(3-hydroxybutyrate) (PHB) and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) [3-5]. Recently, halophilic Archaea have been attracting strong interest in PHBV resear