Bioprocess for hydrolysis of galacto-oligosaccharides in soy molasses and tofu whey by recombinant pseudomonas chlororaphis

Authors: Solaiman, Daniel; Ashby, Richard - Rick; Cross, Nicole

Submitted to: Biocatalysis and Agricultural Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/4/2020
Publication Date: 3/5/2020
Publication URL: https://handle.nal.usda.gov/10113/6842774
Citation: Solaiman, D., Ashby, R.D., Crocker, N.V. 2020. Bioprocess for hydrolysis of galacto-oligosaccharides in soy molasses and tofu whey by recombinant pseudomonas chlororaphis. Biocatalysis and Agricultural Biotechnology. 24:101529. https://doi.org/10.1016/j.bcab.2020.101529.
DOI: https://doi.org/10.1016/j.bcab.2020.101529

Interpretive Summary: We previously discovered a nonharmful microorganism called Pseudomonas chlororaphis (Pc) capable of producing biodetergent (or surfactant) and bioplastic. On the other hand, a low- or no-cost soybean-processing byproduct called soy molasses (SM) is potentially an attractive feedstock to grow Pc. We had previously genetically engineered (GE) Pc to enable it to make enzyme to degrade the chemically complex sugars in SM, and then showed that the GE-Pc could breakdown the soy sugars only when first treated with a chemical called EDTA to make the cells leaky. In the present study, we constructed recombinant enzymes carrying signal sequence to be transported to outside the cells. We also developed a high-cell-density bioprocess without using EDTA to test the degradation of soy sugars by GE-Pc. Results showed that a soy sugar called raffinose was effectively degraded by GE-Pc either in pure chemical form or as a component in a crude SM medium. Another soy sugar called stachyose was only minimally degraded. We believe that continued research on the GE-Pc strains and the bioprocess will eventually allow us to use SM as a low-cost feedstock for production of bioproducts.

Technical Abstract: Objective of the study was to develop bioprocess to hydrolyse galacto-oligosaccharides using genetically engineered Pseudomonas chlororaphis which could produce biopolymer and biosurfactant. To this end alpha-galactosidase genes were expressed in P. chlororaphis transformants either via a plasmid vector or integrated into the chromosome. Furthermore, pseudomonas secretion signals for protein were tested to facilitate extracellular secretion of the gene product. A two-stage fermentation process was then developed to enable hydrolysis of galacto-oligosaccharide by the P. chlororaphis recombinants. The outcome were recombinant P. chlororaphis strains shown to effectively hydrolyse raffinose (a galacto-oligosaccharide) and metabolise the released galactose product.