On the substrate specificity of the rice strigolactone biosynthesis enzyme DWARF27.

ABSTRACT

MAIN CONCLUSION: The ß-carotene isomerase OsDWARF27 is stereo- and double bond-specific. It converts bicyclic carotenoids with at least one unsubstituted ß-ionone ring. OsDWARF27 may contribute to the formation of α-carotene-based strigolactone-like compounds. Strigolactones (SLs) are synthesized from all-trans-ß-carotene via a pathway involving the ß-carotene isomerase DWARF27, the carotenoid cleavage dioxygenases 7 and 8 (CCD7, CCD8), and cytochrome P450 enzymes from the 711 clade (MAX1 in Arabidopsis). The rice enzyme DWARF27 was shown to catalyze the reversible isomerization of all-trans- into 9-cis-ß-carotene in vitro. ß-carotene occurs in different cis-isomeric forms, and plants accumulate other carotenoids, which may be substrates of DWARF27. Here, we investigated the stereo and substrate specificity of the rice enzyme DWARF27 in carotenoid-accumulating E. coli strains and in in vitro assays performed with heterologously expressed and purified enzyme. Our results suggest that OsDWARF27 is strictly double bond-specific, solely targeting the C9-C10 double bond. OsDWARF27 did not introduce a 9-cis-double bond in 13-cis- or 15-cis-ß-carotene. Substrates isomerized by OsDWARF27 are bicyclic carotenoids, including ß-, α-carotene and ß,ß-cryptoxanthin, that contain at least one unsubstituted ß-ionone ring. Accordingly, OsDWARF27 did not produce the abscisic acid precursors 9-cis-violaxanthin or -neoxanthin from the corresponding all-trans-isomers, excluding a direct role in the formation of this carotenoid derived hormone. The conversion of all-trans-α-carotene yielded two different isomers, including 9'-cis-α-carotene that might be the precursor of strigolactones with an ε-ionone ring, such as the recently identified heliolactone.