Abstract
We designed and constructed a green, sustainable, and nongenetically
modified organism (non-GMO) bioprocess to efficiently coproduce
D-tagatose, bioethanol, and microbial protein from whey
powder. First, a one-pot biosynthesis process involving lactose
hydrolysis and D-galactose redox reactions for
D-tagatose production was established in vitro via a
three-enzyme cascade. Second, a nicotinamide adenine dinucleotide
phosphate-dependent galactitol dehydrogenase mutant, D36A/I37R, based on
the nicotinamide adenine dinucleotide-dependent polyol dehydrogenase
from Paracoccus denitrificans was created through rational design and
screening. Moreover, an NADPH recycling module was created in the
oxidoreductive pathway, and the tagatose yield increased by 3.35-fold
compared with that achieved through the pathway without the cofactor
cycle. The reaction process was accelerated using an enzyme assembly
with a glycine–serine linker, and the tagatose production rate was
9.28-fold higher than the initial yield. Finally, Saccharomyces
cerevisiae was introduced into the reaction solution, and 266.5 g of
D-tagatose, 371.3 g of bioethanol, and 215.4 g of dry
yeast (including 38% protein) were obtained from 1 kg of whey powder
(including 810 g lactose). This study provides a promising non-GMO
process for functional food (D-tagatose) production.
Moreover, this process fully utilized whey powder, demonstrating good
atom economy.