Expanding the Portfolio by a Novel Monomeric Oleate Hydratase from Pediococcus parvulus

Abstract

Oleate hydratases convert oleic acid into 10‐hydroxy stearic acid, a valuable fine chemical, useful in lubricant and surfactant formulations. They are of large interest due to their high expression rates and solubility, however, they differ drastically by their overall stability and pH‐ and temperature ranges. To expand their portfolio, another oleate hydratase named OhyPp (originating from Pediococcus parvulus) was characterized. It is a close relative of the well‐known oleate hydratase OhyRe from Rhodococcus erythropolis. OhyPp is only the second member of the monomeric oleate hydratase family with some surprising catalytic features. A distinct characteristic is OhyPp's higher affinity towards FAD compared to OhyRe's helping to understand and improve FAD binding in the future, which is a current drawback for the industrial application of oleate hydratases.

Two Cascade Reactions with Oleate Hydratases for the Sustainable Biosynthesis of Fatty Acid-Derived Fine Chemicals

“…Additionally, the composition of the triglycerides might affect the yield due to the preferences of the lipase [42,43]. Linseed oil contains large amounts of C18:2 and C18:3, which are also substrates for OhyRe [17] and other OHs [18], and thus, could bind and might lead to side reactions, resulting in a lower yield, whereas in C. oleaginosus oil, the most abundant fatty acid after oleic acid is C16:0 (Table 1). Thus, this oil might be beneficial regarding the oleic acid content relative to other C18 species.…”

“…Surprisingly, the first pH optimum shifted towards a more basic environment compared to when 12-HSA was used as the substrate. Since oleic acid has a pk a of 5.02 [18], we investigated the effect on the pH and whether this might be the reason for the activity shift. We indeed observed that the pH was shifted when 12-HSA was present in the reaction buffer, but it could not explain the change in the optimum (Table S1), thus indicating that the type of substrate for secADH can influence the reaction optima.…”

“…The broad-range temperature optimum is beneficial for a one-pot reaction; however, the high pH optimum of secADH is problematic since the optimum of most OHs is rather in the acidic range, presumably due to an evolutionary adaptation to the acidic nature of the substrates. OhyPp [18] and OhyEm [16], for instance, have an optimum of 6. OhyRe, however, is an exception since it prefers rather basic pH environments [17].…”

“…Upon cell lysis, FADH 2 is oxidized to FAD, and in many OHs, only low FAD occupancy can be measured after protein purification, thus making the direct proof of FADH 2 binding preferences considerably challenging. For instance, OhyRe [17], OH from Pediococcus parvulus (OhyPp) [18], and a linoleate hydratase (LAH) [19] have been observed to fully lose FAD upon purification [18,19].…”

“…Several OHs with varying properties are already described and characterized [17,18,24,25], however, monomeric OHs exhibit the greatest potential for an industrial application, as subunit dissociation can be neglected and immobilization strategies can be adopted easier [26]. Hence, this study focuses on an OH from Rhodococcus erythropolis (OhyRe) as the first identified member of monomeric OHs.…”