Biosynthesis of Polyhydroxyalkanoates From Sucrose by Recombinant Pseudomonas putida KT2440

Abstract

A sucrose-utilization pathway was developed in Pseudomonas putida using sacC from Mannheimia succiniciproducens, which encodes a β-fructofuranosidase that hydrolyzes sucrose into glucose and fructose. Excretion of β-fructofuranosidase into the culture medium was confirmed via western blot analysis. In nitrogen-limited cultivation, P. putida expressing SacC produced 10.52 wt % medium-chain-length polyhydroxyalkanoate (MCL-PHA), while P. putida expressing SacC along with poly(3-hydroxybutyrate) [P(3HB)] biosynthesis genes produced 9.16 wt % P(3HB) from sucrose. Batch and fed-batch cultures of recombinant P. putida suggested that the glucose and fructose derived from sucrose can be completely utilized for cell growth and P(3HB) production. In fed-batch cultures, sucrose supplied into the fermentor to maintain its concentration around 20 g/L was rapidly hydrolyzed into glucose and fructose supporting the production of 30.2 g/L P(3HB) with 38.1 wt %. The engineered P. putida reported herein can facilitate the production of PHAs from sucrose, an abundant and inexpensive carbon source.