Structure-based protein engineering enables prenyl donor switching of a fungal aromatic prenyltransferase

Mai, Peter; Zocher, Georg; Stehle, Thilo; Li, Shu-Ming

doi:10.1039/c8ob02037j

Cited by 19 publications

(37 citation statements)

References 33 publications

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“…Simultaneously, the K M value of DMAPP was almost stable in all enzyme synthesis reactions of substrates. This result also proved that DMAPP acted as a stable isopentenyl donor in the prenylation reaction [ 32 , 53 ]. The order of K M , k c at and k c at / K M for seven substrates was inconsistent.…”

Section: Resultsmentioning

confidence: 58%

“…It has been hypothesized speculated that due to the increase in the hydrophobicity of 2,5-DKPs by adding isoprene groups, the prenylated molecules are better distributed on the membrane than the non-prenylated molecules and are more closely related to the target protein [ 8 , 9 , 35 , 53 ]. In the structures ( 1b – 7b ), the prenyl moieties were connected via its C1 to indole ring C7.…”

Section: Resultsmentioning

confidence: 99%

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C7-Prenylation of Tryptophan-Containing Cyclic Dipeptides by 7-Dimethylallyl Tryptophan Synthase Significantly Increases the Anticancer and Antimicrobial Activities

Zhang

et al. 2020

Molecules

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Prenylated natural products have interesting pharmacological properties and prenylation reactions play crucial roles in controlling the activities of biomolecules. They are difficult to synthesize chemically, but enzymatic synthesis production is a desirable pathway. Cyclic dipeptide prenyltransferase catalyzes the regioselective Friedel–Crafts alkylation of tryptophan-containing cyclic dipeptides. This class of enzymes, which belongs to the dimethylallyl tryptophan synthase superfamily, is known to be flexible to aromatic prenyl receptors, while mostly retaining its typical regioselectivity. In this study, seven tryptophan-containing cyclic dipeptides 1a–7a were converted to their C7-regularly prenylated derivatives 1b–7b in the presence of dimethylallyl diphosphate (DMAPP) by using the purified 7-dimethylallyl tryptophan synthase (7-DMATS) as catalyst. The HPLC analysis of the incubation mixture and the NMR analysis of the separated products showed that the stereochemical structure of the substrate had a great influence on their acceptance by 7-DMATS. Determination of the kinetic parameters proved that cyclo-l-Trp–Gly (1a) consisting of a tryptophanyl and glycine was accepted as the best substrate with a KM value of 169.7 μM and a turnover number of 0.1307 s−1. Furthermore, docking studies simulated the prenyl transfer reaction of 7-DMATS and it could be concluded that the highest affinity between 7-DMATS and 1a. Preliminary results have been clearly shown that prenylation at C7 led to a significant increase of the anticancer and antimicrobial activities of the prenylated derivatives 1b–7b in all the activity test experiment, especially the prenylated product 4b.

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Section: Resultsmentioning

confidence: 58%

Section: Resultsmentioning

confidence: 99%

C7-Prenylation of Tryptophan-Containing Cyclic Dipeptides by 7-Dimethylallyl Tryptophan Synthase Significantly Increases the Anticancer and Antimicrobial Activities

Zhang

et al. 2020

Molecules

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“…The geranylation of tryptophan was only reported for two 6-DMATS from Streptomyces violaceusniger and Streptomyces ambofaciens that both produced 6-geranyl tryptophan . Fungal DMATS usually do not accept GPP instead of DMAPP, but lately enzyme variants of the 4-DMATS FgaPT2 and several diketopiperazine (DKP) converting DMATS obtained by site directed mutagenesis were shown to produce geranylated products. The only occurrence of reverse N -geranylation on the indole nucleus was reported as one of four products from the incubation of a F335G mutant of CdpC3PT with cyclo-Trp-Val .…”

Section: Resultsmentioning

confidence: 99%

Biochemical and Mechanistic Characterization of the Fungal Reverse N-1-Dimethylallyltryptophan Synthase DMATS1_Ff

Burkhardt

Janevska

et al. 2019

ACS Chem. Biol.

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Dimethylallyltryptophan synthases catalyze the regiospecific transfer of (oligo)prenylpyrophosphates to aromatic substrates like tryptophan derivatives. These reactions are key steps in many biosynthetic pathways of fungal and bacterial secondary metabolites. In vitro investigations on recombinant DMATS1 Ff from Fusarium f ujikuroi identified the enzyme as the first selective reverse tryptophan-N-1 prenyltransferase of fungal origin. The enzyme was also able to catalyze the reverse N-geranylation of tryptophan. DMATS1 Ff was shown to be phylogenetically related to fungal tyrosine O-prenyltransferases and fungal 7-DMATS. Like these enzymes, DMATS1 Ff was able to convert tyrosine into its regularly O-prenylated derivative. Investigation of the binding sites of DMATS1 Ff by homology modeling and comparison to the crystal structure of 4-DMATS FgaPT2 showed an almost identical site for DMAPP binding but different residues for tryptophan coordination. Several putative active site residues were verified by site directed mutagenesis of DMATS1 Ff . Homology models of the phylogenetically related enzymes showed similar tryptophan binding residues, pointing to a unified substrate binding orientation of tryptophan and DMAPP, which is distinct from that in FgaPT2. Isotopic labeling experiments showed the reaction catalyzed by DMATS1 Ff to be nonstereospecific. Based on these data, a detailed mechanism for DMATS1 Ff catalysis is proposed.

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“…The substitution with smaller side chain, including M328A, M328C, M328S, and M328G significantly increased the activity for GPP and FPP prenylation (Fig. 5d) [107]. Furthermore, the model also suggested that the active site residues Lue263 and Tyr398 could also interfere with terminal isoprene unit of FPP.…”

Section: Structure-based Engineering Of Abba and Dmats Ptasesmentioning

confidence: 89%

Enzymatic studies on aromatic prenyltransferases

Mori

2020

J Nat Med

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Aromatic prenyltransferases (PTases), including ABBA-type and dimethylallyl tryptophan synthase (DMATS)-type enzymes from bacteria and fungi, play important role for diversification of the natural products and improvement of the biological activities. For a decade, the characterization of enzymes and enzymatic synthesis of prenylated compounds by using ABBA-type and DMATS-type PTases have been demonstrated. Here, I introduce several examples of the studies on chemoenzymatic synthesis of unnatural prenylated compounds and the enzyme engineering of ABBA-type and DMATS-type PTases.

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Structure-based protein engineering enables prenyl donor switching of a fungal aromatic prenyltransferase

Abstract: Structure-guided molecular modelling and site-directed mutagenesis of the tryptophan dimethylallyl transferase FgaPT2 led to creation of mutants with strongly enhanced activities towards geranyl and farnesyl diphosphates.

Cited by 19 publications

References 33 publications

C7-Prenylation of Tryptophan-Containing Cyclic Dipeptides by 7-Dimethylallyl Tryptophan Synthase Significantly Increases the Anticancer and Antimicrobial Activities

C7-Prenylation of Tryptophan-Containing Cyclic Dipeptides by 7-Dimethylallyl Tryptophan Synthase Significantly Increases the Anticancer and Antimicrobial Activities

Biochemical and Mechanistic Characterization of the Fungal Reverse N-1-Dimethylallyltryptophan Synthase DMATS1_Ff

Enzymatic studies on aromatic prenyltransferases

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Structure-based protein engineering enables prenyl donor switching of a fungal aromatic prenyltransferase

Abstract: Structure-guided molecular modelling and site-directed mutagenesis of the tryptophan dimethylallyl transferase FgaPT2 led to creation of mutants with strongly enhanced activities towards geranyl and farnesyl diphosphates.

Cited by 19 publications

References 33 publications

C7-Prenylation of Tryptophan-Containing Cyclic Dipeptides by 7-Dimethylallyl Tryptophan Synthase Significantly Increases the Anticancer and Antimicrobial Activities

C7-Prenylation of Tryptophan-Containing Cyclic Dipeptides by 7-Dimethylallyl Tryptophan Synthase Significantly Increases the Anticancer and Antimicrobial Activities

Biochemical and Mechanistic Characterization of the Fungal Reverse N-1-Dimethylallyltryptophan Synthase DMATS1Ff

Enzymatic studies on aromatic prenyltransferases

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Biochemical and Mechanistic Characterization of the Fungal Reverse N-1-Dimethylallyltryptophan Synthase DMATS1_Ff