Premium
Poly(3‐hydroxybutyrate‐ co ‐3‐hydroxyhexanoate) Production in Recombinant Aeromonas hydrophila 4AK4 Harboring phbA , phbB and vgb Genes
Author(s) -
Ouyang Shaoping,
Han Jing,
Qiu Yuanzheng,
Qin Lingfang,
Chen Song,
Wu Qiong,
Leski Michael L.,
Chen Guoqiang
Publication year - 2005
Publication title -
macromolecular symposia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.257
H-Index - 76
eISSN - 1521-3900
pISSN - 1022-1360
DOI - 10.1002/masy.200550603
Subject(s) - aeromonas hydrophila , strain (injury) , recombinant dna , chemistry , cell growth , bacteria , microbiology and biotechnology , biology , biochemistry , gene , genetics , anatomy
Poly(3‐hydroxybutyrate‐ co ‐3‐hydroxyhexanoate) (PHBHHx), a copolyester consisting of 3‐hydroxybutyrate (3HB) and 3‐hydroxyhexanoate (3HHx), can be synthesized by Aeromonas hydrophila strain 4AK4 using long chain fatty acids as the carbon source. The wild type A. hydrophila 4AK4 accumulated PHBHHx consisting of 12‐15 mol% 3HHx regardless of growth conditions. When phbA , phbB and vgb genes encoding β ‐ketothiolase, acetoacetyl‐CoA reductase and vitreoscilla hemoglobin, respectively, were introduced together into A. hydrophila 4AK4, the recombinant strain grew to over 20 g/L cell dry weight (CDW) after 48 h of shake flask cultivation in co‐substrates of dodecanoate and gluconate (weight ratio 1:1), and the CDW contained 50% PHBHHx consisting of 9 mol% 3HHx. Under similar conditions, the wild type strain produced only 12 g/L CDW containing 32% PHBHHx with 15 mol% 3HHx. In comparison, recombinant harboring phbA and phbB produced 35% PHBHHx with 9 mol% 3HHx in 15 g/L CDW under the same conditions. The obvious differences in terms of the cell growth and PHBHHx production were attributed to the expression of vgb in A. hydrophila 4AK4, which was clearly observed in carbon monoxide difference spectra. The expression of vgb in the recombinant not only improved cell growth and PHBHHx accumulation, but also increased the plasmid stability during cell growth, especially under low dissolved oxygen tension in fermentors. PHBHHx production could be further increased to over 60% of the CDW by the over expression of phaC and phaJ from Aeromonas caviae encoding PHBHHx synthase and (R)‐specific enoyl‐CoA hydratase, respectively. Over expression of phaC , phaJ and phaP , alone or in various combinations, also increased the 3HHx content of PHBHHx from 14‐34%. The above results showed that A. hydrophila was amenable to genetic manipulation, and that these modifications could be exploited to produce compounds with different properties for commercial and research applications.