Biosynthesis of blasticidin S from cytosylglucuronic acid (CGA). Isolation of cytosine/UDPglucuronosyltransferase and incorporation of CGA by Streptomyces griseochromogenes

Guo, Jincan; Gould, Steven J.

doi:10.1021/ja00015a074

Cited by 20 publications

(15 citation statements)

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“…Classical radiolabeled precursor studies established the primary precursors as glucose, cytosine, L ‐arginine, and methionine 26. Through the use of both radioactive and stable isotope‐labeled compounds, β ‐arginine, cytosylglucuronic acid (CGA), and cytosinine were identified as advanced intermediates in the pathway 16, 27, 28. Efforts to unravel the biochemical steps in the assembly of 1 resulted in the purification and characterization of CGA synthase, the enzyme catalyzing the first committed step in the pathway, the condensation of cytosine and UDP‐glucuronic acid (UDP, uridine diphosphate) to form CGA 15.…”

Section: Introductionmentioning

confidence: 99%

The Blasticidin S Biosynthesis Gene Cluster from Streptomyces griseochromogenes: Sequence Analysis, Organization, and Initial Characterization

et al. 2003

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Blasticidin S is a potent antifungal and cytotoxic peptidyl nucleoside antibiotic from Streptomyces griseochromogenes. The mixed biosynthesis of the compound is evident from the three distinct structural components: a cytosine base, an amino deoxyglucuronic acid, and N-methyl beta-arginine. The blasticidin S biosynthesis gene cluster was cloned from S. griseochromogenes and the pathway heterologously expressed in S. lividans from a cosmid harboring a 36.7-kb fragment of S. griseochromogenes DNA. The complete DNA sequence of this insert has now been determined and evidence suggests a contiguous 20-kb section defines the blasticidin S biosynthesis cluster. The predicted functions of several open reading frames are consistent with the expected biochemistry and include an arginine 2,3-aminomutase, a cytosylglucuronic acid synthase, and a guanidino N-methyltransferase. Insight into other steps in the assembly of blasticidin S was evident from sequence homology with proteins of known function and heterologous expression of fragments of the cluster. Additionally, the gene that directs the production of free cytosine, blsM, was subcloned and expressed in Escherichia coli. Characterization of BlsM revealed that cytidine monophosphate serves as the precursor to cytosine.

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Section: Introductionmentioning

confidence: 99%

The Blasticidin S Biosynthesis Gene Cluster from Streptomyces griseochromogenes: Sequence Analysis, Organization, and Initial Characterization

et al. 2003

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“…1) (14), and N-demethylblasticidin S (12). We have established that CGA is formed from cytosine (2 in Fig.…”

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confidence: 99%

“…As shown in Figure 2, SDS-PAGE of the purified CGA synthase from the last step (lanes 8 CGA synthase, isolated from S. griseochromogenes, catalyzes the formation of CGA from UDP-glucuronic acid and cytosine in the biosynthesis of the peptide-nucleoside antibiotic BS (14) and is also quite likely involved in the biosynthesis of a number of other known nucleoside antibiotics (1,7,15,19,20,36,37). Functionally, CGA synthase belongs to the family of UDPGTs; although UDP-GTs are common in mammalian metabolism and at least two have been reported for fungi (10,40), they have not been previously reported for a prokaryotic organism.…”

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confidence: 99%

Cytosylglucuronic acid synthase (cytosine: UDP-glucuronosyltransferase) from Streptomyces griseochromogenes, the first prokaryotic UDP-glucuronosyltransferase

Gould

Guo

1994

J Bacteriol

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Cytosylglucuronic acid synthase (cytosine: UDP-glucuronosyltransferase), the first prokaryotic UDP-GT and a key enzyme in the biosynthesis of the antibiotic blasticidin S, was purified 870-fold. It has optimum activity at a pH of 8.4 to 8.6, Kms of 6.0 (UDP-glucuronic acid) and 243 (cytosine) microM, and a maximum rate of metabolism of 14.6 mumol/min/mg. The apparent M(r) is 43,000. Activity was slightly enhanced by Mg2+ or Ca2+ but was not inhibited by EDTA. Activity was strongly inhibited by UDP. Cytosylglucuronic acid differs from eukaryotic UDP-glucuronosyltransferases in being a soluble protein with no apparent phospholipid requirement.

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“…S7), indicating that P10 is a nucleoside antibiotic similar to blasticidin S (compound 1) (13)(14)(15)(16). Owing to the different 13 C shifts for C-10 in compound 2 compared to compound 1 and to the presence of an NH versus an O atom, compound 2 was determined to be the amide form of blasticidin S. The absolute configuration of blasticidin S (compound 1) has been reported previously from chemical degradation and X-ray crystallography experiments (17,18). The nearly iden-tical NMR spectra (Fig.…”

Section: Resultsmentioning

confidence: 67%

A New Natural Product Analog of Blasticidin S Reveals Cellular Uptake Facilitated by the NorA Multidrug Transporter

Davison

Lohith

Wang

et al. 2017

Antimicrob Agents Chemother

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The permeation of antibiotics through bacterial membranes to their target site is a crucial determinant of drug activity but in many cases remains poorly understood. During screening efforts to discover new broad-spectrum antibiotic compounds from marine sponge samples, we identified a new analog of the peptidyl nucleoside antibiotic blasticidin S that exhibited up to 16-fold-improved potency against a range of laboratory and clinical bacterial strains which we named P10. Whole-genome sequencing of laboratory-evolved strains of Staphylococcus aureus resistant to blasticidin S and P10, combined with genome-wide assessment of the fitness of barcoded Escherichia coli knockout strains in the presence of the antibiotics, revealed that restriction of cellular access was a key feature in the development of resistance to this class of drug. In particular, the gene encoding the well-characterized multidrug efflux pump NorA was found to be mutated in 69% of all S. aureus isolates resistant to blasticidin S or P10. Unexpectedly, resistance was associated with inactivation of norA, suggesting that the NorA transporter facilitates cellular entry of peptidyl nucleosides in addition to its known role in the efflux of diverse compounds, including fluoroquinolone antibiotics.

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Biosynthesis of blasticidin S from cytosylglucuronic acid (CGA). Isolation of cytosine/UDPglucuronosyltransferase and incorporation of CGA by Streptomyces griseochromogenes

Cited by 20 publications

References 0 publications

The Blasticidin S Biosynthesis Gene Cluster from Streptomyces griseochromogenes: Sequence Analysis, Organization, and Initial Characterization

The Blasticidin S Biosynthesis Gene Cluster from Streptomyces griseochromogenes: Sequence Analysis, Organization, and Initial Characterization

Cytosylglucuronic acid synthase (cytosine: UDP-glucuronosyltransferase) from Streptomyces griseochromogenes, the first prokaryotic UDP-glucuronosyltransferase

A New Natural Product Analog of Blasticidin S Reveals Cellular Uptake Facilitated by the NorA Multidrug Transporter

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