Biosynthesis of antibiotics of the virginiamycin family. 8. Formation of the dehydroproline residue

Purvis, Michael B.; LeFevre, Joseph W.; Jones, Vickie L.; Kingston, David G. I.; Biot, A.; Gosselé, F.

doi:10.1021/ja00197a064

Cited by 18 publications

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“…that produce pristinamycins. The same type of observation was made with Streptomyces virginiae, the producer of virginiamycin, closely related to pristinamycin (49). These results indicated that PII B is the biosynthetic precursor of PII A , and so the oxidation of the proline residue into a dehydroproline residue appears to be the last step of PII A biosynthesis.…”

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confidence: 65%

Cloning and analysis of structural genes from Streptomyces pristinaespiralis encoding enzymes involved in the conversion of pristinamycin IIB to pristinamycin IIA (PIIA): PIIA synthase and NADH:riboflavin 5'-phosphate oxidoreductase

Blanc¹,

Lagneaux²,

Didier³

et al. 1995

J Bacteriol

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. 177:5199-5205, 1995); these enzymes are PII A synthase, a heterodimer composed of the SnaA and SnaB proteins, which catalyzes the oxidation of PII B to PII A , and the NADH:riboflavin 5-phosphate oxidoreductase (hereafter called FMN reductase), the SnaC protein, which provides the reduced form of flavin mononucleotide for the reaction. By using oligonucleotide probes designed from limited peptide sequence information of the purified proteins, the corresponding genes were cloned from a genomic library of S. pristinaespiralis. SnaA and SnaB showed no significant similarity with proteins from databases, but SnaA and SnaB had similar protein domains. Disruption of the snaA gene in S. pristinaespiralis led to accumulation of PII B . Complementation of a S. pristinaespiralis PII A ؊ PII B ؉ mutant with the snaA and snaB genes, cloned in a low-copy-number plasmid, partially restored production of PII A . The deduced amino acid sequence of the snaC gene showed no similarity to the sequences of other FMN reductases but was 39% identical with the product of the actVB gene of the actinorhodin cluster of Streptomyces coelicolor A(3)2, likely to be involved in the dimerization step of actinorhodin biosynthesis. Furthermore, an S. coelicolor A(3)2 mutant blocked in this step was successfully complemented by the snaC gene, restoring the production of actinorhodin.Pristinamycin belongs to the family of streptogramin antibiotics, also called virginiamycin-like or mikamycin-like antibiotics. Streptogramins are a small and homogeneous group composed of related compounds such as pristinamycin, virginiamycin, mikamycin, and vernamycin (9, 10, 58). They are protein synthesis inhibitors (9, 10). The special feature of the family is that each member is a complex of two structurally different components exhibiting a synergistic antibacterial activity (2, 10). The two types of compounds are both macrocyclic lactone peptolides, but their structures are notably different. They belong to one of the two following distinct groups: the streptogramin A type (Sa) corresponding to polyunsaturated cyclic peptolides and the streptogramin B type (Sb) corresponding to branched cyclic hexadepsipeptides. The proportion of Sa and Sb in the complex varies from one antibiotic to another. Moreover, the major form of each component is accompanied by several structurally different minor forms.Pristinamycin, produced by Streptomyces pristinaespiralis, consists of approximately 30% pristinamycins I (PI), the Sb type molecules, and 70% pristinamycins II (PII), the Sa type molecules. In industrial strains, PII is produced mainly in two forms, PII A and PII B , in a 80:20 ratio. The difference between PII A and PII B is the presence of a dehydroproline instead of a proline in the macrocycle (Fig. 1). Thibaut et al. (57) reported high levels of conversion of radiolabelled PII B to PII A both in vivo and in vitro with several strains of Streptomyces spp. that produce pristinamycins. The same type of observation was made with Streptomyces virginiae, the producer ...

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supporting

confidence: 65%

Cloning and analysis of structural genes from Streptomyces pristinaespiralis encoding enzymes involved in the conversion of pristinamycin IIB to pristinamycin IIA (PIIA): PIIA synthase and NADH:riboflavin 5'-phosphate oxidoreductase

Blanc¹,

Lagneaux²,

Didier³

et al. 1995

J Bacteriol

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“…In the presence of PII B , about one molecule of oxygen is consumed to produce one molecule of PII A , indicating that PII B is first hydroxylated. Because the 3-R proton of the D-proline residue of PII B is lost during the formation of PII A (25), hydroxylation probably occurs at this position (reaction 2 in Fig. 5).…”

Section: Discussionmentioning

confidence: 99%

“…Purvis and coworkers tried to elucidate the mechanism of formation of the unique dehydroproline residue contained in PII A and replaced by a D-proline in PII B . By using stereospecifically labelled L-and D-proline, they showed that both L-and D-proline are precursors of the dehydroproline residue and that the 3-R proton of L-proline is still present in PII B , while it is stereospecifically lost in PII A (25). This established that D-proline cannot be derived from dehydroproline, and as a result, PII A cannot be a precursor of PII B .…”

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confidence: 99%

“…Conversely, weak conversion of radiolabelled PII B to PII A was obtained, favoring the hypothesis that PII B lies on the pathway to PII A . Moreover, incorporation of cis/trans hydroxyproline into the dehydroproline residue of PII A suggested that dehydration is likely to generate the double bond (25).…”

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confidence: 99%

“…The concentrations used were 0.15 to 1 M for FMN, 0 to 15 M for lumichrome, and 500 M for NADH. (25). Because PII A production seemed to occur earlier in this strain, another feeding experiment was done with the addition of [ 14 C]PII B at the beginning of fermentation in the production medium.…”

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confidence: 99%

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Purification of the two-enzyme system catalyzing the oxidation of the D-proline residue of pristinamycin IIB during the last step of pristinamycin IIA biosynthesis

et al. 1995

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High levels of conversion of 14 C-labelled pristinamycin II B (PII B ) to pristinamycin II A (PII A ) were obtained in vivo in Streptomyces pristinaespiralis and in some other streptogramin A producers. This established that PII B was an intermediate on the pathway to PII A . In addition, in vitro studies with cell-free protein preparations demonstrated that the oxidation of PII B to PII A is a complex process requiring NADH, riboflavin 5-phosphate (FMN), and molecular oxygen. Two enzymes were shown to be necessary to catalyze this reaction. Both were purified to homogeneity from S. pristinaespiralis by a coupled enzyme assay based on the formation of PII A and by requiring addition of the complementing enzyme. One enzyme was purified about 3,000-fold by a procedure including a decisive affinity chromatography step on FMN-agarose. It was shown to be a NADH:FMN oxidoreductase (E.C. 1.6.8.1.) (hereafter called FMN reductase), providing reduced FMN (FMNH 2 ) to the more abundant second enzyme. The latter was purified only 160-fold and was called PII A synthase. Our data strongly suggest that this enzyme catalyzes a transient hydroxylation of PII B by molecular oxygen immediately followed by a dehydration leading to PII A . The native PII A synthase consists of two different subunits with M r s of around 50,000 and 35,000, as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, while the FMN reductase seems to be a monomer with a M r of around 28,000 and containing one molecule of tightly bound FMN. Stepwise Edman degradation of the entire polypeptides or some of their trypsin-digested fragments provided amino acid sequences for the two isolated proteins.Pristinamycins I and pristinamycins II (PII), the two families of bacteriostatic components associated in the bactericidal antibiotic pristinamycin, are produced by Streptomyces pristinaespiralis (22,23). Pristinamycins I are cyclohexadepsipeptides belonging to the B group of streptogramins, while PII are polyunsaturated macrolactones of the A group of streptogramins (Fig. 1). Both streptogramins A and B are secondary metabolites synthesized by a large number of different Streptomycetes, and most of the members of these families have received several names, often related to the species from which they were isolated (6, 28). Thus, the main component of PII, pristinamycin II A (PII A ), is also called mikamycin A, ostreogrycin A, staphylomycin M1, streptogramin A, synergistin A, vernamycin A, virginiamycin M1, and PA-114 A1, while the minor component pristinamycin II B (PII B ) is also known as ostreogrycin G and virginiamycin M2 (7).Pristinamycin is poorly water soluble, and its therapeutic use has remained rather restricted until now. The development of a water-soluble streptogramin, RP 59500, has recently revived interest in this old family of antibiotics (1). Pristinamycin is active against both gram-positive bacteria, especially Staphylococcus and Streptococcus spp., and some gram-negative bacteria, such as Haemophilus spp. Interestingly, both ...

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Determination of enantiomeric purity of commercial 14C- and 3H-labeled L-α-amino acids

LeFevre

Bonzagni

Chappell

et al. 1998

J Label Compd Radiopharm

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The enantiomeric purity of twelve commercial 14C‐ and 3H‐labeled L‐α‐amino acids was determined using reverse isotope dilution analysis. The technique utilized reversed‐phase (RP) thin‐layer chromatography (TLC) and beta‐cyclodextrin (β‐CD) in the mobile phase to separate D‐ and L‐amino acids as their 5‐dimethylamino‐1‐naphtalene sulfonyl (dansyl, DNS) derivatives. In all cases, the L‐amino acid was contaminated with the D‐isomer. This is the first report of the resolution of N‐DNS‐Dl‐tyrosine and N‐(α)‐DNS‐Dl‐lysine using this methodology. © 1998 John Wiley & Sons, Ltd.

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Biosynthesis of antibiotics of the virginiamycin family. 8. Formation of the dehydroproline residue

Cited by 18 publications

References 0 publications

Cloning and analysis of structural genes from Streptomyces pristinaespiralis encoding enzymes involved in the conversion of pristinamycin IIB to pristinamycin IIA (PIIA): PIIA synthase and NADH:riboflavin 5'-phosphate oxidoreductase

Cloning and analysis of structural genes from Streptomyces pristinaespiralis encoding enzymes involved in the conversion of pristinamycin IIB to pristinamycin IIA (PIIA): PIIA synthase and NADH:riboflavin 5'-phosphate oxidoreductase

Purification of the two-enzyme system catalyzing the oxidation of the D-proline residue of pristinamycin IIB during the last step of pristinamycin IIA biosynthesis

Determination of enantiomeric purity of commercial 14C- and 3H-labeled L-α-amino acids

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