example) and a carboxylate ending chain should be place somewhere on the hydro-
carbon chain of the substrate. To fulfill these two requirements, we used methyl
15(S)-HETE previously synthesized from SBLOX-1 and AA in the normal orienta-
tion followed by hydroperoxide reduction and esterification (Martini et al., 1996a).
Then, by taking advantage of the 15-hydroxy functionality, we added a succinyl
chain (from succinic anhydride) to methyl 15(S)-HETE. Thus the newly synthesized
substrate 9 should be recognized by SBLOX-1 in such a way that the formal 5-oxy-
genation of the carbon skeleton of AA will be allowed (Figure 12).
Compound 9 is indeed a substrate of SBLOX-1, but should be used at low sub-
strate concentration (10-3M) since at higher substrate concentrations the 5-hydro-
peroxide formed is unstable in the reaction medium. Nevertheless, by using a 2-
L fermentor, the reaction could be conducted on hundreds of milligram scale,
and it was shown that the second dioxygenation performed by SBLOX-1 was essen-
tially diastereoselective. Thus, this work represents a chemo-enzymatic synthesis of
the naturally occurring 5(S),15(S)-diHETE which is obtained in 59 % yields and
more than 98 % diastereoisomeric excess (Martini et al., 1996a).
16.3 Substrate and product specificities of lipoxygenases 349
Figure 12. Synthesis of a derivative of AA prompt to be oxygenated at the formal 5 position by
SBLOX-1 (Martini et al., 1996a).