Biotechnological Production of Odor-Active Methyl-Branched Aldehydes by a Novel α-Dioxygenase from Crocosphaera subtropica

Abstract

As a result of their pleasant odor qualities and low odor thresholds, iso-and anteiso-fatty aldehydes represent promising candidates for applications in flavoring preparations. A novel cyanobacterial α-dioxygenase from Crocosphaera subtropica was heterologously expressed in Escherichia coli and applied for the biotechnological production of C 12 −C 15 branched-chain fatty aldehydes. The enzyme has a sequence identity of less than 40% to well-investigated α-dioxygenase from rice. Contrary to the latter, it efficiently transformed short-chained fatty acids. The kinetic parameters of α-dioxygenase toward unbranched and iso-branchedchain substrates were studied by means of an oxygen-depletion assay. The transformation products (C 12 −C 15 iso-and anteisoaldehydes) were extensively characterized, including their sensory properties. The aldehydes exhibited green-soapy, sweety odors with partial citrus-like, metallic, peppery, and savory-tallowy nuances. Moreover, the two C 14 isomers showed particularly low odor threshold values of 0.2 and 0.3 ng/L in air as determined by means of gas chromatography−olfactometry.

Two novel cyanobacterial α-dioxygenases for the biosynthesis of fatty aldehydes

“…2 ). From these results, the cyanobacterial α-DOXs newly identified in our study were found to be functionally equivalent to plant α-DOXs in terms of their heme-dependency, which was not elucidated for the previously described cyanobacterial CsDOX (Hammer et al 2020 ).

Fig.

…”

“…Although most α-DOXs studied so far are from plants (Kaehne et al 2011;Koeduka et al 2005;Liu et al 2006), the existence of α-DOX enzymes in non-plant organisms should not be excluded considering the recent discovery of a cyanobacterial α-DOX (Hammer et al 2020). However, the complex phylogeny across taxa had not been examined After incubating for the indicated times, the biotransformations were stopped by the addition of 2-M HCl (30 μL), and the samples were extracted three times with an equal volume of ethyl acetate yet.…”

“…They occur in a wide range of living organisms including plants, insects, and mammals as metabolic intermediates of lipids (Foster and Anderson 2019 ; Gilbertson et al 1972 ; Liu et al 2012 ). Many types of fatty aldehydes provide pleasant and diverse odor impressions depending on their chemical structure determined by chain length, the presence/number/position of double bonds, and further substitutions (Hammer et al 2020 ). Thus, they are widely applied in the food, perfume, cosmetic, and detergent industries as important flavor and fragrance compounds.…”

“…However, issues related to technical complexity and expensive process cost cannot be avoided when using this enzyme class as they require the cofactors ATP, NADPH, and Mg 2+ and the additional expression of a partner enzyme, a phosphopantetheinyl transferase (PPTase) (Akhtar et al 2013 ; Maurer et al 2019 ; Wu et al 2021 ). In contrast, α-dioxygenases (α-DOXs), a family of heme-dependent fatty acid dioxygenases that convert FFAs into fatty aldehydes, can be potent candidates to tackle these problems as they require only molecular oxygen, but no expensive cofactors (Goulah et al 2013 ; Hamberg et al 2002 ; Hammer et al 2020 ; Kaehne et al 2011 ; Koeduka et al 2005 ).…”

“…Although this fatty acid α-oxidation mechanism is well-known in plants and, accordingly, catalytic functions of various α-DOXs from plants have been thoroughly elucidated, there is only one functional study of an α-DOX from a non-plant organism, the CsDOX from the cyanobacterium Crocosphaera subtropica (Hammer et al 2020 ). CsDOX was revealed to have a catalytically equivalent function to α-DOX from plants but shows a different substrate specificity.…”