Manipulation of prenylation reactions by structure-based engineering of bacterial indolactam prenyltransferases

Takahiro Mori,Masahiro Okada,Ikuro Abe,Takayoshi Awakawa,Shotaro Hoshino,Hiroyuki Morita,Lihan Zhang

doi:10.1038/ncomms10849

Takahiro Mori, Masahiro Okada + Show 5 more

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https://doi.org/10.1038/ncomms10849

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Abstract

Prenylation reactions play crucial roles in controlling the activities of biomolecules. Bacterial prenyltransferases, TleC from Streptomyces blastmyceticus and MpnD from Marinactinospora thermotolerans, catalyse the ‘reverse' prenylation of (−)-indolactam V at the C-7 position of the indole ring with geranyl pyrophosphate or dimethylallyl pyrophosphate, to produce lyngbyatoxin or pendolmycin, respectively. Using in vitro analyses, here we show that both TleC and MpnD exhibit relaxed substrate specificities and accept various chain lengths (C5–C25) of the prenyl donors. Comparisons of the crystal structures and their ternary complexes with (−)-indolactam V and dimethylallyl S-thiophosphate revealed the intimate structural details of the enzyme-catalysed ‘reverse' prenylation reactions and identified the active-site residues governing the selection of the substrates. Furthermore, structure-based enzyme engineering successfully altered the preference for the prenyl chain length of the substrates, as well as the regio- and stereo-selectivities of the prenylation reactions, to produce a series of unnatural novel indolactams.

Highlights

Prenylation reactions play crucial roles in controlling the activities of biomolecules
As prenylated molecules usually exhibit improved interactions with proteins and biological membranes, prenylation reactions catalysed by prenyltransferases (PTs) play crucial roles in controlling the activities of biomolecules, as exemplified by the biosyntheses of Vitamin K and menaquinones in primary metabolism[1], the posttranslational modifications of peptides[2,3] and proteins[4], as well as the production of various biologically active secondary metabolites in plants, fungi and bacteria[5,6,7,8,9]
We recently reported the biosynthetic gene cluster for teleocidin B, a unique prenylated indolactam with a monoterpenoid moiety fused with C-6 and C-7 of the indole ring[14] (Fig. 1a)

Summary

Introduction

Prenylation reactions play crucial roles in controlling the activities of biomolecules. As prenylated molecules usually exhibit improved interactions with proteins and biological membranes, prenylation reactions catalysed by prenyltransferases (PTs) play crucial roles in controlling the activities of biomolecules, as exemplified by the biosyntheses of Vitamin K and menaquinones in primary metabolism[1], the posttranslational modifications of peptides[2,3] and proteins[4], as well as the production of various biologically active secondary metabolites in plants, fungi and bacteria[5,6,7,8,9] This is the case for the indole alkaloids, which include the specific cell cycle inhibitors tryprostatins[5], the ergot alkaloids anti-migraine drugs[6] and the potent protein kinase C activator teleocidins[7] (Fig. 1). We succeeded in structure-based enzyme engineering, to rationally control the preference for the chain lengths of the prenyl donors, as well as the regio- and stereo-selectivities of the enzyme reactions, to generate unnatural novel indolactams

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Conclusion