4.2.1

Cofactor Dependence

The production of xylitol and L-arabitol during pentose consumption by natural as well as recombinant pentose-utilizing yeasts has been rational­ized with the difference in cofactor preferences between the enzymes in the initial pentose utilization pathways. XR from P stipitis preferentially uses NADPH, but can also use NADH as a cofactor [23], whereas XDH exclu­sively uses NAD+ [24,46]. This may result in excess NADH formation and lack of NAD+, since yeasts do not harbor a transhydrogenase enzyme that would allow direct conversion of NADP+ to NAD+ [32]. Numerous investiga­tions have supported this metabolic model; external electron acceptors, which are reduced by NADH-dependent enzymes in S. cerevisiae, reduce xylitol for­mation [32,74-76]. Xylitol formation in recombinant S. cerevisiae was also reduced by changing the kinetic properties of the enzymes involved by ex­pressing a fusion protein of XR and XDH [87], or by expressing mutated XR with altered cofactor affinity [88,89] (strains TMB3270, TMB3271, R267H, Ta­bles 1-3). However, significantly increased ethanolic xylose fermentation as a result of such engineering strategies has been less frequently reported [89].

4.2.2