Crystal structures of a 6-dimethylallyltryptophan synthase, IptA Insights into substrate tolerance and enhancement of prenyltransferase activity

Dimethylallyltryptophan synthases (DMATSs) catalyze the prenyl transfer reaction from dimethylallyl pyrophosphate (DMAPP) to an indole ring. IptA, a member of the DMATS family, is involved in biosyn -thesis of 6-dimethylallylindole-3-carbaldehyde in Streptomyces sp. SN-593 and catalyzes the C6-prenylation of L-Trp. The enzyme exhibits prenyl acceptor promiscuity and can accept various Trp de-rivatives, as observed in several other DMATS family members. Although many crystal structures of DMATS have been determined to date, the structural basis of substrate promiscuity and the acceptance of alternatives to indole-containing natural substrates remain to be clarified. In this study, we determined the crystal structures of the ternary L-Trp derivative (5-methyl-, 6-methyl-, and Na-methyl-L-Trp)-DMSPP (dimethylallyl S-thiolopyrophosphate; stable analog of DMAPP)-enzyme complex of IptA, in addition to the substrate-free IptA and ternary L-Trp-DMSPP-IptA complex crystal structures. The overall structure of IptA exhibited a typical ABBA-fold, which is commonly found in DMATS family members, while L-Trp and DMSPP are found in a tunnel located inside the ABBA barrel. The crystal structure of the ternary L-Trp-DMSPP-enzyme complex can explain the electrophilic substitution at the C6 atom of L-Trp, which is assisted by Glu84 and His294, as previously suggested for other DMATSs. Although L-Trp snugly fitted into the active site pocket and the unoccupied space around L-Trp is very limited in the L-Trp-DMSPP-IptA complex structure, the enzyme can accommodate 5 -methyl-and 6-methyl-L-Trp by slight relocation of the substrate indole ring and adjacent side chain in the active site, resulting in a higher prenylation activity for 5-methyl-L-Trp and C7 prenylation of 6-methyl-L-Trp. Like many other DMATSs, IptA cannot utilize prenyl donors larger than DMAPP. To enlarge the prenyl donor-binding pocket, the W154A mutation was introduced. As expected, this mutant produced prenylated L-Trp from L-Trp and geranyl-and farnesyl pyrophosphate. (c) 2022 Elsevier Inc. All rights reserved.

Automated summary

In this study, the crystal structures of IptA were determined, providing insights into the substrate selectivity and prenyl acceptor promiscuity. The study found that IptA can accommodate different L-Trp derivatives by adjusting the position of the substrate indole ring and adjacent side chain, resulting in different prenylation activities.