Tertiary alcohol preferred: Hydroxylation of trans-3-methyl-L-proline with proline hydroxylases

Abstract

The enzymatic synthesis of tertiary alcohols by the stereospecific oxidation of tertiary alkyl centers is a most-straightforward but challenging approach, since these positions are sterically hindered. In contrast to P450-monooxygenases, there is little known about the potential of non-heme iron(II) oxygenases to catalyze such reactions. We have studied the hydroxylation of trans-3-methyl-L-proline with the α-ketoglutarate (α-KG) dependent oxygenases, cis-3-proline hydroxylase type II and cis-4-proline hydroxylase (cis-P3H_II and cis-P4H). With cis-P3H_II, the tertiary alcohol product (3R)-3-hydroxy-3-methyl-L-proline was obtained exclusively but in reduced yield (~7%) compared to the native substrate L-proline. For cis-P4H, a complete shift in regioselectivity from C-4 to C-3 was observed so that the same product as with cis-P3H_II was obtained. Moreover, the yields were at least as good as in control reactions with L-proline (~110% relative yield). This result demonstrates a remarkable potential of non-heme iron(II) oxygenases to oxidize substrates selectively at sterically hindered positions.

Keywords

• asymmetric catalysis
• enzyme catalysis
• hydroxyproline
• α-ketoglutarate dependent iron(II) oxygenases
• regioselectivity
• stereoselectivity