Biocatalytic synthesis of non-vicinal aliphatic diols

Abstract

Regioselective biocatalytic oxyfunctionalization of n-alkanes for the production of non-vicinal diols through sequential oxygenation by a cytochrome P450 monooxygenase.

Structure of the fungal hydroxylase, CYP505A30, and rational transfer of mutation data from CYP102A1 to alter regioselectivity

“…1.0 M for n-decane biotransformations. [b] TTN and TOF values in parenthesis were previously reported for CYP505A30wt by Ebrecht et al 22…”

“…CYP102A1 produces a majority of ω-3 (C4) hydroxylated products from n-octane and n-decane, whereas CYP505A30's regioselectivity favours hydroxylation at the ω-1 (C2) and ω-2 (C3) positions (even though regioselectivity is also dependent on substrate length). 22,29 We anticipated these differences in regioselectivity to arise due to variations in amino acids in the active site. Comparing the active sites of both enzymes, we noticed two distinct alanine residues in close proximity to the haem in CYP102A1 (A82 and A328) that are replaced by valine (V88) and isoleucine (I334), respectively, in CYP505A30 (Figure 2).…”

“…The achieved turnover numbers of CYP505A30wt are, however, lower than the activities we have reported previously (reduction by ca. 13 -27 %) 22 , which can be due to the different conditions we used in this set of experiments (e.g. lower temperature of 20 °C vs 30 °C, Tris buffer vs potassium phosphate buffer).…”

“…19 Diol formation was already detected in the biotransformation reaction after 1 h. The total concentration of diols produced by CYP505A30 reactions was on average 5.6 mM of octanediol and 5.3 mM of decanediol after 24 h. Overall, the relative percentage of diols to all products formed in n-decane biotransformations was higher than in n-octane biotransformations (Figure S12), possibly due to the low solubility of n-decane compared to the higher solubility of the n-decanol products. 22 Control reactions with CYP102A1 showed that it also produces diols but to a lesser percentage of overall products formed (Figure S14). Please do not adjust margins Please do not adjust margins A similar pattern of diol formation was observed for both noctane and n-decane biotransformations.…”

“…21 We recently reported an in-depth characterisation of the products produced by CYP505A30 from Myceliophthora thermophila. 22 This self-sufficient CYP450 formed a mixture of non-vicinal diols through sequential hydroxylation of n-octane and n-decane. Here, we report the X-ray crystal structure of the haem domain of CYP505A30.…”