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Research Call

DAFM Reference

Lead(Collaborating)Institution

DAFM Award

DAFM National Call 2015 15F747 NUIG (Teagasc) €98,877

Project Title:

Enzymes for efficient milk oligosaccharide production (EFFICIenz)

Project Coordinator:

Prof Lokesh Joshi

Project Abstract

This application builds upon an ongoing FIRM-funded project “Enzymatic generation of sialylated lactose from waste whey using marine-derived sialyltransferases” (SIALenz). Two strategies were proposed for the production of sialylated oligosaccharides for use in infant formula supplementation. One strand described in vitro reaction of sialic acid and lactose catalysed by novel animal sialyltransferases while the second strand described bacterial fermentation on lactose using a metabolically engineered Lactococcus strain. The enzymatic addition of sialic acid (Neu5Ac) to lactose cannot be achieved efficiently and with specificity by chemical synthesis. A number of studies have shown that the sialylated oligosaccharides, possess anti-adhesive effects against certain pathogens and pro-adhesive effects for beneficial commensal strains. Sialyllactoses are present in bovine milk but at much lower levels than in human making isolation from bovine milk an impractical option at present.

A commercial necessity for the production of sialyllactoses by the in vitro approach using novel purified CHO expressed sialyltransferases is an inexpensive supply of the substrate CMP-Neu5Ac (CMP-activated sialic acid). We propose to use purified CMP synthetase (CMAS) to convert Neu5Ac to CMP-Neu5Ac. Because CMP-Neu5Ac is relatively unstable, a secondary goal is to fuse CMAS with ST6Gal1 making a complex that transfers labile CMP-Neu5Ac directly from CMAS to ST6Gal1 thereby making 6’sialyllactose from lactose and sialic acid. In our second approach to HMO production, we propose to use the expertise gained in the SIALenz project to expand our repertoire of oligosaccharides to the production of fucosylated oligosaccharides. Fucosylated HMOs are important in shaping the infant gut microbiome and are associated with a lower risk of diarrhoea and respiratory disease. We are currently in the final stages of creating a metabolically engineered E. coli strain capable of producing 3’ and 6’-sialyllactose. We will use a similar recombineering approach and enzymes derived from marine bacteria/non-human pathogen sources to produce 2’fucosyllactose

Final Report:

Not available yet.