#Inmr normalize integrals free

The biotechnological production of versatile chemicals in a sustainable manner depends on the availability of low-cost biomass and efficient microbial production hosts. While the conversion efficiency of the process can be further improved by enzyme engineering, the reaction demonstrates the first multi-enzyme cascade for the bioconversion of SCG. Finally, 4.4 ± 0.1 mM lactic acid was produced from 14.57 ± 0.7 mM SCG-derived mannose. Moreover, reactions with isotope labeled mannose enabled the monitoring of pathway intermediates and revealed a reverse flux in the conversion process. Assessment of enzymes, adjustment of enzyme loadings, substrate and cofactor concentrations, and buffer ionic strength allowed the identification of crucial reaction parameters and bottlenecks.

#Inmr normalize integrals free

In comparison to known in vivo metabolisms the bioconversion is free of phosphorylated intermediates and cofactors. The enzymatic conversion is based on a so far unknown oxidative mannose metabolism from Thermoplasma acidophilum and uses a previously characterized mannonate dehydratase to convert mannose into lactic acid via 4 enzymatic reactions. Here, we present a cell-free reaction cascade for the conversion of mannose, the most abundant sugar in SCG, into L-lactic acid. Spent coffee grounds (SCG) are an industrially underutilized waste resource, which is rich in the polysaccharide galactomannan, a polysaccharide consisting of a mannose backbone with galactose side groups. The coffee industry produces over 10 billion kg beans per year and generates high amounts of different waste products. 2Biomolecular Discovery and Design Research Centre, Macquarie University, Sydney, NSW, Australia.1Department of Molecular Sciences, Macquarie University, Sydney, NSW, Australia.