Regiospecific Benzylation of Tryptophan and Derivatives Catalyzed by a Fungal Dimethylallyl Transferase

A big challenge in organic synthesis is to reach a high regioselectivity. Enzymes catalyze usually highly regiospecific reactions and can function as ideal biocatalysts for such purposes. Some secondary metabolite enzymes can even use distinctly different unnatural substrates and expand therefore their potential usage in chemoenzymatic synthesis. We report here the acceptance of benzyl diphosphate as an alkyl donor by the fungal dimethylallyl transferase FgaPT2 and the regiospecific enzymatic benzylation of tryptophan and several analogues

Mutations of Residues in Pocket P1 of a Cyclodipeptide Synthase Strongly Increase Product Formation

Expression of a cyclodipeptide synthase gene from <i>Nocardiopsis prasina</i> (<i>CDPS-Np</i>) in <i>Escherichia coli</i> resulted in the formation of <i>cyclo</i>-(l-Tyr-l-Tyr) (<b>1</b>) as the minor and <i>cyclo</i>-(l-Tyr-l-Phe) (<b>2</b>) as the major products. Site-directed mutagenesis revealed a strong influence on product accumulation of the amino acid residues in pocket P1. An 8-fold increase in product formation for <b>1</b> and 10-fold for <b>2</b> were detected in the double mutant T82V_Y196F compared with the wild type

Breaking Cyclic Dipeptide Prenyltransferase Regioselectivity by Unnatural Alkyl Donors

The behavior of five cyclic dipeptide prenyltransferases, responsible for C2-regular, C2-reverse, or C3-reverse prenylation, was investigated in the presence of the unnatural alkyl donors monomethylallyl and 2-pentenyl diphosphate. Both substrates were well accepted by the tested enzymes. Interestingly, C2-reverse and C3-reverse monoalkylated derivatives were identified as enzyme products in all of the enzyme assays. These findings indicate their similar reaction characteristics in the presence of unnatural alkyl donors

Expansion of Enzymatic Friedel–Crafts Alkylation on Indoles: Acceptance of Unnatural β‑Unsaturated Allyl Diphospates by Dimethylallyl-tryptophan Synthases

Prenyltransferases of the dimethylallyl-tryptophan synthase (DMATS) superfamily catalyze Friedel–Crafts alkylation with high flexibility for aromatic substrates, but the high specificity for dimethylallyl diphosphate (DMAPP) prohibits their application as biocatalysts. We demonstrate here that at least one methyl group in DMAPP can be deleted or shifted to the δ-position. For acceptance by some DMATS enzymes, however, a double bond must be situated at the β-position. Furthermore, the alkylation position of an analogue can differ from that of DMAPP

Coupling of Guanine with <i>cyclo</i>-l‑Trp‑l‑Trp Mediated by a Cytochrome P450 Homologue from <i>Streptomyces purpureus</i>

A cyclo-l-Trp-l-Trp tailoring P450 with novel function from <i>Streptomyces purpureus</i> was identified by heterologous expression in <i>S. coelicolor</i> and in vitro assays the recombinant protein. Structural elucidation revealed that this enzyme catalyzes the transfer of a guanine moiety to the indole ring of the cyclodipeptide via a C–N bond. Adduct products of CDP and guanine are unprecedented in nature, and CDP modification by coupling with guanine has not been reported prior to this study

Friedel–Crafts Alkylation of Acylphloroglucinols Catalyzed by a Fungal Indole Prenyltransferase

Naturally occurring prenylated acylphloroglucinol derivatives are plant metabolites with diverse biological and pharmacological activities. Prenylation of acylphloroglucinols plays an important role in the formation of these intriguing natural products and is catalyzed in plants by membrane-bound enzymes. In this study, we demonstrate the prenylation of such compounds by a soluble fungal prenyltransferase AnaPT involved in the biosynthesis of prenylated indole alkaloids. The observed activities of AnaPT toward these substrates are much higher than that of a microsomal fraction containing an overproduced prenyltransferase from the plant hop

Formation of the Fungal Indole Alkaloid Speradine F Implies Multiple Nonenzymatic Oxidation Steps

The highly oxygenated indole alkaloid speradine F (4) with a 6/5/6/5/5/5 hexacyclic skeleton was isolated from a culture of Penicillium palitans, together with its precursors β-cyclopiazonic acid (β-CPA, 5) and cyclopiazonic acid (CPA, 1). Gene deletion and heterologous expression led to the identification of the responsible five-gene spe cluster for the speradine skeleton formation. Precursor supply experiments proved that 1 was enzymatically converted, via 2-oxoCPA (2), to speradine A (3), which subsequently undergoes multistep nonenzymatic hydroxylations to 4

A <i>Streptomyces</i> Cytochrome P450 Enzyme Catalyzes Regiospecific <i>C</i>2‑Guaninylation for the Synthesis of Diverse Guanitrypmycin Analogs

Heterologous expression of a cdps-p450 locus from Streptomyces sp. NRRL S-1521 led to the identification of guanitrypmycin D1, a new guaninylated diketopiperazine. The cytochrome P450 GutD1521 catalyzed the regiospecific transfer of guanine to C-2 of the indole ring of cyclo-(l-Trp-l-Tyr) via a C–C linkage and represents a new chemical transformation within this enzyme class. Furthermore, GutD1521 efficiently accepts several other tryptophan-containing cyclodipeptides or derivatives for regiospecific coupling with guanine, thus generating different guanitrypmycin analogs

Complementary Flavonoid Prenylations by Fungal Indole Prenyltransferases

Flavonoids are found mainly in plants and exhibit diverse biological and pharmacological activities, which can often be enhanced by prenylations. In plants, such reactions are catalyzed by membrane-bound prenyltransferases. In this study, the prenylation of nine flavonoids from different classes by a soluble fungal prenyltransferase (AnaPT) involved in the biosynthesis of the prenylated indole alkaloid acetylaszonalenin is demonstrated. The behavior of AnaPT toward flavonoids regarding substrate acceptance and prenylation positions clearly differs from that of the indole prenyltransferase 7-DMATS. The two enzymes are therefore complementary in flavonoid prenylations