Stereospecific Biosynthesis of β‐Methyltryptophan from <scp>L</scp>‐Tryptophan Features a Stereochemical Switch

Zou, Yi; Yin, Haixing; Liang, Zutao; Kong, Dekun; Bai, Linquan; Deng, Zixin; Lin, Shuangjun

doi:10.1002/ange.201306255

Cited by 15 publications

(12 citation statements)

References 39 publications

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“…Subsequently, StnK3 catalyzes epimerization, and StnR catalyzes a second round of transamination to complete the assembly of (2S,3S)-β-methyltryptophan. 22,23 To confirm the role of stnR in streptonigrin biosynthesis, stnR was inactivated in vivo by replacing an internal fragment of stnR with the apramycin resistance cassette to generate WJN-1, and the genotype was verified by PCR (Supplementary Figure S1). To evaluate the effect of stnR replacement, the production of streptonigrin in WJN-1 was monitored by HPLC and LC-MS. Streptonigrin production was completely abolished in the strain WJN-1 and, surprisingly, two new compounds (1 and 2), absent in the wild-type strain, were present in the mutant strain WJN-1 ( Figure 2 and Supplementary Figure S16).…”

Section: Resultsmentioning

confidence: 99%

Xantholipin B produced by the stnR inactivation mutant Streptomyces flocculus CGMCC 4.1223 WJN-1

Huang

Xie

et al. 2016

J Antibiot

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Xantholipin is a polycyclic xanthone antibiotic that exhibits potent cytotoxic and antibacterial activity. In this study, a new xanthone-type antibiotic, xantholipin B (1), was isolated for the first time along with its known derivative, xantholipin (2), from strain WJN-1, an aminotransferase inactivation mutant of the streptonigrin-producer Streptomyces flocculus CGMCC 4.1223. The structure of 1 was established based on spectroscopic analysis and supports the previously proposed biosynthetic pathway as a key intermediate of 2. Moreover, 1 showed 3- to 10-fold greater cytotoxicity than 2 against a select panel of human cancer cell lines. In addition, 1 demonstrated powerful antimicrobial activity against both Gram-positive bacteria and fungi. Importantly, both 1 and 2 inhibited the methicillin-resistant strain Staphylococcus aureus Mu50, with the MIC value of 0.025 μg ml. The new structural features of 1 enrich the structural diversity of xantholipin family compounds and shed new light on the structure-activity relationship of 1 as a promising antitumor drug candidate.

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Section: Resultsmentioning

confidence: 99%

Xantholipin B produced by the stnR inactivation mutant Streptomyces flocculus CGMCC 4.1223 WJN-1

Huang

Xie

et al. 2016

J Antibiot

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“…[21] For example, C-MTases that catalyse the stereoselective β-methylation of α-ketoacid substrates are of interest, and in combination with aminotransferase enzymes provide access to a range of non-proteinogenic β-methyl-α-amino acids. [17,[44][45][46] Many synthetic peptidomimetics and other pharmaceuticals are derived from β-methyl-α-amino acid building blocks, which are currently prepared using expensive multi-step synthetic processes. It is envisaged that the α-ketoacid C-MTases could be used in combination with aminotransferases to provide a more efficient and cleaner route to β-methyl-α-amino acids and derivatives.…”

Section: )mentioning

confidence: 99%

Recent advances in methyltransferase biocatalysis

Bennett

Shepherd

Cronin

et al. 2017

Current Opinion in Chemical Biology

102

111

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S-adenosyl-L-methionine-dependent methyltransferses are ubiquitous in nature, methylating a vast range of small molecule metabolites, as well as biopolymers. This review covers the recent advances in the development of methyltransferase enzymes for synthetic applications, focusing on the methyltransferase catalyzed transformations with S-adenosyl methionine analogs, as well as non-native substrates. We discuss how metabolic engineering approaches have been used to enhance S-adenosyl methionine production in vivo. Enzymatic approaches that enable the more efficient generation of S-adenosyl methionine analogs, including more stable analogs, will also be described; this has expanded the biocatalytic repertoire of methyltransferases from methylation to a broader range of alkylation reactions. The review also examines how the selectivity of the methyltransferase enzymes can be improved through structure guided mutagenesis approaches. Finally, we will discuss how methyltransferases can be deployed in multi-enzyme cascade reactions and suggest future challenges and avenues for further investigation.

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“…In order to characterize the enzymes of this cassette in vitro , the three genes were individually overexpressed in E. coli BL21(DE3) and the corresponding N -terminally His 6 -tagged proteins were purified to near homogeneity ( Figure S2 ). Subsequently, the enzymatic activities of these three proteins were evaluated following a previously described procedure 34 . StnR catalyzed the transamination of l -Trp to generate indolepyruvate when it was incubated with the amino acceptor α-ketoglutarate ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Bioinformatic analysis of the stn gene cluster revealed a three-gene cassette ( stnQ1-stnK3-stnR ) encoding an S -adenosylmethionine (SAM)-dependent C -methyltransferase, a cupin-fold epimerase, and a pyridoxal 5′-phosphate (PLP)-dependent aminotransferase, respectively. This gene cassette is homologous to the marI-marH-marG cassette that generates (2 S ,3 S )-β-MeTrp for the biosynthetic pathway of maremycins ( Figure S1 ; identity/similarity: 64%/77% for StnQ1/MarI, 82%/86% for StnK3/MarH, and 64%/73% for StnR/MarG) 34 . The bioinformatic data suggests that the stnQ1-stnK3-stnR cassette is responsible for the biosynthesis of (2 S ,3 S )-β-MeTrp as well.…”

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Identification of (2S,3S)-β-Methyltryptophan as the Real Biosynthetic Intermediate of Antitumor Agent Streptonigrin

Kong

Zou

Zhang

et al. 2016

Sci Rep

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Streptonigrin is a potent antitumor antibiotic, active against a wide range of mammalian tumor cells. It was reported that its biosynthesis relies on (2S,3R)-β-methyltryptophan as an intermediate. In this study, the biosynthesis of (2S,3R)-β-methyltryptophan and its isomer (2S,3S)-β-methyltryptophan by enzymes from the streptonigrin biosynthetic pathway is demonstrated. StnR is a pyridoxal 5′-phosphate (PLP)-dependent aminotransferase that catalyzes a transamination between L-tryptophan and β-methyl indolepyruvate. StnQ1 is an S-adenosylmethionine (SAM)-dependent C-methyltransferase and catalyzes β-methylation of indolepyruvate to generate (R)-β-methyl indolepyruvate. Although StnR exhibited a significant preference for (S)-β-methyl indolepyruvate over the (R)-epimer, StnQ1 and StnR together catalyze (2S,3R)-β-methyltryptophan formation from L-tryptophan. StnK3 is a cupin superfamily protein responsible for conversion of (R)-β-methyl indolepyruvate to its (S)-epimer and enables (2S,3S)-β-methyltryptophan biosynthesis from L-tryptophan when combined with StnQ1 and StnR. Most importantly, (2S,3S)-β-methyltryptophan was established as the biosynthetic intermediate of the streptonigrin pathway by feeding experiments with a knockout mutant, contradicting the previous proposal that stated (2S,3R)-β-methyltryptophan as the intermediate. These data set the stage for the complete elucidation of the streptonigrin biosynthetic pathway, which would unlock the potential of creating new streptonigrin analogues by genetic manipulation of the biosynthetic machinery.

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Stereospecific Biosynthesis of β‐Methyltryptophan from L‐Tryptophan Features a Stereochemical Switch

Cited by 15 publications

References 39 publications

Xantholipin B produced by the stnR inactivation mutant Streptomyces flocculus CGMCC 4.1223 WJN-1

Xantholipin B produced by the stnR inactivation mutant Streptomyces flocculus CGMCC 4.1223 WJN-1

Recent advances in methyltransferase biocatalysis

Identification of (2S,3S)-β-Methyltryptophan as the Real Biosynthetic Intermediate of Antitumor Agent Streptonigrin

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