Controlled autolysis facilitates the polyhydroxyalkanoate recovery in Pseudomonas putida KT2440
Article first published online: 21 MAR 2011
Published 2011. This article is a US Government work and is in the public domain in the USA. Journal compilation © 2011 Society for Applied Microbiology and Blackwell Publishing Ltd
Thematic Issue: Extremophiles
Volume 4, Issue 4, pages 533–547, July 2011
How to Cite
Martínez, V., García, P., García, J. L. and Prieto, M. A. (2011), Controlled autolysis facilitates the polyhydroxyalkanoate recovery in Pseudomonas putida KT2440. Microbial Biotechnology, 4: 533–547. doi: 10.1111/j.1751-7915.2011.00257.x
- Issue published online: 6 JUL 2011
- Article first published online: 21 MAR 2011
- Received 11 August, 2010; accepted 22 November, 2010.
The development of efficient recovery processes is essential to reduce the cost of polyhydroxyalkanoates (PHAs) production. In this work, a programmed self-disruptive Pseudomonas putida BXHL strain, derived from the prototype medium-chain-length PHA producer bacterium P. putida KT2440, was constructed as a proof of concept for exploring the possibility to control and facilitate the release of PHA granules to the extracellular medium. The new autolytic cell disruption system is based on two simultaneous strategies: the coordinated action of two proteins from the pneumococcal bacteriophage EJ-1, an endolysin (Ejl) and a holin (Ejh), and the mutation of the tolB gene, which exhibits alterations in outer membrane integrity that induce lysis hypersensitivity. The ejl and ejh coding genes were expressed under a XylS/Pm monocopy expression system inserted into the chromosome of the tolB mutant strain, in the presence of 3-methylbenzoate as inducer molecule. Our results demonstrate that the intracellular presence of PHA granules confers resistance to cell envelope. Conditions to control the cell autolysis in P. putida BXHL in terms of optimal fermentation, PHA content and PHA recovery have been set up by exploring the sensitivity to detergents, chelating agents and wet biomass solubility in organic solvents such as ethyl acetate.