Bioproduction of P-Hydroxybenzoate From Renewable Feedstock by Solvent-Tolerant Pseudomonas Putida S12

J Biotechnol. 2007 Oct 15;132(1):49-56. doi: 10.1016/j.jbiotec.2007.08.031. Epub 2007 Aug 23.

Abstract

Pseudomonas putida strain S12palB1 was constructed that produces p-hydroxybenzoate from renewable carbon sources via the central metabolite l-tyrosine. P. putida S12palB1 was based on the platform strain P. putida S12TPL3, which has an optimised carbon flux towards l-tyrosine. Phenylalanine ammonia lyase (Pal) was introduced for the conversion of l-tyrosine into p-coumarate, which is further converted into p-hydroxybenzoate by endogenous enzymes. p-Hydroxybenzoate hydroxylase (PobA) was inactivated to prevent the degradation of p-hydroxybenzoate. These modifications resulted in stable accumulation of p-hydroxybenzoate at a yield of 11% (C-molC-mol(-1)) on glucose or on glycerol in shake flask cultures. In a glycerol-limited fed-batch fermentation, a final p-hydroxybenzoate concentration of 12.9mM (1.8gl(-1)) was obtained, at a yield of 8.5% (C-molC-mol(-1)). A 2-fold increase of the specific p-hydroxybenzoate production rate (q(p)) was observed when l-tyrosine was supplied to a steady-state C-limited chemostat culture of P. putida S12palB1. This implied that l-tyrosine availability was the bottleneck for p-hydroxybenzoate production under these conditions. When p-coumarate was added instead, q(p) increased by a factor 4.7, indicating that Pal activity is the limiting factor when sufficient l-tyrosine is available. Thus, two major leads for further improvement of the p-hydroxybenzoate production by P. putida S12palB1 were identified.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Bioreactors
  • Biotechnology
  • DNA Primers / genetics
  • DNA, Bacterial / genetics
  • Fermentation
  • Kinetics
  • Parabens / metabolism*
  • Phenylalanine Ammonia-Lyase / metabolism
  • Plasmids / genetics
  • Pseudomonas putida / genetics
  • Pseudomonas putida / metabolism*
  • Solvents
  • Tyrosine / metabolism

Substances

  • DNA Primers
  • DNA, Bacterial
  • Parabens
  • Solvents
  • Tyrosine
  • Phenylalanine Ammonia-Lyase
  • 4-hydroxybenzoic acid