Molecular characterisation of phaCAB from Comamonas sp. EB172 for functional expression in Escherichia coli JM109.

Autor: Yee LN; Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia., Chuah JA, Chong ML, Phang LY, Raha AR, Sudesh K, Hassan MA
Jazyk: angličtina
Zdroj: Microbiological research [Microbiol Res] 2012 Oct 12; Vol. 167 (9), pp. 550-7. Date of Electronic Publication: 2012 Jan 27.
DOI: 10.1016/j.micres.2011.12.006
Abstrakt: In this study, PHA biosynthesis operon of Comamonas sp. EB172, an acid-tolerant strain, consisting of three genes encoding acetyl-CoA acetyltransferase (phaA(Co) gene, 1182 bp), acetoacetyl-CoA reductase (phaB(Co) gene, 738 bp) and PHA synthase, class I (phaC(Co) gene, 1694 bp) were identified. Sequence analysis of the phaA(Co), phaB(Co) and phaC(Co) genes revealed that they shared more than 85%, 89% and 69% identity, respectively, with orthologues from Delftia acidovorans SPH-1 and Acidovorax ebreus TPSY. The PHA biosynthesis genes (phaC(Co) and phaAB(Co)) were successfully cloned in a heterologous host, Escherichia coli JM109. E. coli JM109 transformants harbouring pGEM'-phaC(Co)AB(Re) and pGEM'-phaC(Re)AB(Co) were shown to be functionally active synthesising 33 wt.% and 17 wt.% of poly(3-hydroxybutyrate) [P(3HB)]. E. coli JM109 transformant harbouring the three genes from the acid-tolerant Comamonas sp. EB172 (phaCAB(Co)) under the control of native promoter from Cupriavidus necator, in vivo polymerised P(3HB) when fed with glucose and volatile mixed organic acids (acetic acid:propionic acid:n-butyric acid) in ration of 3:1:1, respectively. The E. coli JM109 transformant harbouring phaCAB(Co) could accumulate P(3HB) at 2g/L of propionic acid. P(3HB) contents of 40.9% and 43.6% were achieved by using 1% of glucose and mixed organic acids, respectively.
(Copyright © 2012 Elsevier GmbH. All rights reserved.)
Databáze: MEDLINE
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