Molecular basis for differential elongation of omega-3 docosapentaenoic acid by the rat Elovl5 and Elovl2

Abstract

and Elovl2, which have been overlooked as regulators of DHA synthesis. Using a yeast expression system, it was apparent that the substrate specifi cities of the two rat elongases had some overlap, but that only Elovl2 could convert endogenously formed C 22 PUFA docosapentaenoic acid (DPA) (22:5n-3) to 24:5n-3, which is the penultimate precursor of DHA ( 5 ). Elovl2 performs the sequential elongation of EPA to DPA followed by further elongation to 24:5n-3.Thus, Elovl2 is crucial for DHA synthesis at least in the rat where Elovl5 cannot elongate DPA to 24:5n-3 ( 5 ). This probably explains the poor or absent ability to produce DHA in species that do not have detectable Elovl2 such as barramundi ( 6, 7 ) or in species such as the rat in which Elovl2 is expressed at low levels ( 5 ). However, there is not an absolute Elovl2 dependence for DHA synthesis in all species because the sea bream, cobia, Atlantic bluefi n tuna, and chicken Elovl5 have a small but measurable ability to elongate DPA ( 8-11 ). In order to better understand the potential for these elongases to be involved in DHA synthesis, we have sought the molecular reasons for the differences between Elovl5 and Elovl2 in their ability to elongate DPA to 24:5n-3.Purifi cation of membrane-bound elongases to determine the substrate binding pocket has proven to be unsuccessful ( 12 ). However, chimeric elongase proteins from yeast ( 13 ), the moss Physcomitrella patens ( 14 ), and the fungi Pythium irregulare and Phytophthore infestans ( 15 ) have been used to investigate the regions involved in C 18 and C 20 PUFA substrate specifi city and product chain length determination. Therefore, we have constructed a series of rat Elovl2/Elovl5 chimeras and point mutations to examine the Elovl2 residues responsible for DPA substrate specifi city using a yeast expression system.Abstract Functional characterization of the rat elongases, Elovl5 and Elovl2, has identifi ed that Elovl2 is crucial for omega-3 docosahexaenoic acid (DHA) (22:6n-3) synthesis. While the substrate specifi cities of the rat elongases had some overlap, only Elovl2 can convert the C 22 omega-3 PUFA docosapentaenoic acid (DPA) (22:5n-3) to 24:5n-3, which is the penultimate precursor of DHA. In order to better understand the potential for these elongases to be involved in DHA synthesis, we have examined the molecular reasons for the differences between Elovl5 and Elovl2 in their ability to elongate DPA to 24:5n-3. We identifi ed a region of heterogeneity between Elovl5 and Elovl2 spanning transmembrane domains 6 and 7. Using a yeast expression system, we examined a series of Elovl2/Elovl5 chimeras and point mutations to identify Elovl2 residues within this region which are responsible for DPA substrate specifi city. The results indicate that the cysteine at position 217 in Elovl2 and a tryptophan at the equivalent position in Elovl5 explain their differing abilities to elongate DPA to 24:5n-3. Further studies confi rmed that Elovl2 C217 is a critical residue for elongation of DPA at the level observed i...

Molecular basis of fatty acid selectivity in the zDHHC family of S-acyltransferases revealed by click chemistry

“…Elovl2 is required for synthesis of omega-3 docosahexaenoic acid (DHA; 22:6n-3), because this elongase (but not Elovl5) can elongate docosapentaenoic acid (22:5n-3) to 24:5n-3, a precursor of DHA. This difference in substrate specificity between Elovl2 and Elovl5 was shown to involve a region encompassing transmembrane domains 6 and 7, with a cysteine-to-tryptophan switch in transmembrane domain 7 proving particularly important in setting specificity (31).…”

Diversification of substrate specificities in teleostei Fads2: characterization of Δ4 and Δ6Δ5 desaturases of Chirostoma estor

“…Provided the Gnathostomata (jawed fi sh) ancestral Fads2 had ⌬ 6 desaturase activity ( 32 ), as for mammalian FADS2s ( 35 ), the expansion of Fads2s in teleosts has been accompanied by subfunctionalization in the enzyme derived from independent mutations in the primary aa sequence ( 32 ). Although a recent study identifi ed a single aa residue as determining the differential ability for 22:5n-3 elongation between ELOVL2 and ELOVL5 elongases in rat ( 48 ), identifi cation of specifi c domains/residues controlling the functionality of desaturases has been elusive and studies are restricted to nonvertebrate enzymes (49)(50)(51). However, the above said, the existence of three ⌬ 6 ⌬ 5 desaturases and three ⌬ 4 desaturases distributed in four distinct species, allows us to explore potential evolutionary scenarios for teleostei Fads2 subfunctionalization.…”

Meibomian glands, meibum, and meibogenesis

“…The microarray data demonstrated a modest expression of ELOVL2 (averaging 20% in the tarsal plates of mice and 30% – in humans), but high expression levels of the gene in the livers of mice (about 80–85%) and humans (about 60%). The ELOVL2 enzyme catalyzes elongation of C 20 -C 22 polyunsaturated n-6 series FA with three or more double bonds, such as C 20 and C 22 polyunsaturated FA (Gregory et al, 2011); (Gregory et al, 2013a); (Okuda et al, 2010), but is also fairly active with C 14 -C 24 saturated substrates (Kitazawa et al, 2009). The low expression levels of ELOVL2/Elovl2 in TP of both species are consistent with our previous reports on the very small presence of its products in human and mouse meibum (Butovich, 2008; Butovich et al, 2012a; Butovich et al, 2007b; Butovich et al, 2009).…”