The Biosynthesis of Vancomycin-Type Glycopeptide Antibiotics-The Order of the Cyclization Steps

Bischoff, Daniel; Pelzer, Stefan; Bister, Bojan; Nicholson, Graeme; Stockert, Sigrid; Schirle, Markus; Wohlleben, Wolfgang; Jung, Günther; Süßmuth, Roderich D.

doi:10.1002/1521-3773(20011217)40:24<4688::aid-anie4688>3.0.co;2-m

Cited by 137 publications

(96 citation statements)

References 20 publications

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“…In an attempt to obtain a structure of the OxyB substrate complex, OxyB was co-crystallized with the putative substrate SP1066 (1, X ϭ H) (20). Although the peptide was not visible in either crystal form (ASnative 2, PEG-native 2), the data allowed us to build the region Arg …”

Section: Methodsmentioning

confidence: 99%

Crystal Structure of OxyB, a Cytochrome P450 Implicated in an Oxidative Phenol Coupling Reaction during Vancomycin Biosynthesis

Zerbe

Pylypenko

Vitali

et al. 2002

Journal of Biological Chemistry

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134

118

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Gene-inactivation studies point to the involvement of OxyB in catalyzing the first oxidative phenol coupling reaction during glycopeptide antibiotic biosynthesis. The oxyB gene has been cloned and sequenced from the vancomycin producer Amycolatopsis orientalis, and the hemoprotein has been produced in Escherichia coli, crystallized, and its structure determined to 1.7-Å resolution. OxyB gave UV-visible spectra characteristic of a P450-like hemoprotein in the low spin ferric state. After reduction to the ferrous state by dithionite or by spinach ferredoxin and ferredoxin reductase, the CO-ligated form gave a 450-nm peak in a UV-difference spectrum. Addition of putative heptapeptide substrates to resting OxyB produced type I changes to the UV spectrum, but no turnover was observed in the presence of ferredoxin and ferredoxin reductase, showing that either the peptides or the reduction system, or both, are insufficient to support a full catalytic cycle. OxyB exhibits the typical P450-fold, with helix L containing the signature sequence FGHGXHXCLG and Cys 347 being the proximal axial thiolate ligand of the heme iron. The structural similarity of OxyB is highest to P450nor, P450terp, CYP119, and P450eryF. In OxyB, the F and G helices are rotated out of the active site compared with P450nor, resulting in a much more open active site, consistent with the larger size of the presumed heptapeptide substrate.

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Section: Methodsmentioning

confidence: 99%

Crystal Structure of OxyB, a Cytochrome P450 Implicated in an Oxidative Phenol Coupling Reaction during Vancomycin Biosynthesis

Zerbe

Pylypenko

Vitali

et al. 2002

Journal of Biological Chemistry

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134

118

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“…For the vancomycin producer, it has been shown that the homologous oxygenases are P450 monooxygenases (38). OxyA, -B, and -C in A. balhimycina catalyze the cross-linking steps between the aromatic rings within the balhimycin peptide backbone in a defined order (6). In contrast, the function of OxyD remained unclear.…”

Section: Resultsmentioning

confidence: 99%

“…In order to study the biosynthesis of glycopeptide antibiotics and the functions of the relevant genes (6,22,31), we chose the balhimycin producer strain Amycolatopsis balhimycina DSM5908 (36) as a model system. A. balhimycina belongs to the order of Actinomycetales and was formerly described as Amycolatopsis mediterranei (8,18).…”

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confidence: 99%

Biosynthesis of Chloro-β-Hydroxytyrosine, a Nonproteinogenic Amino Acid of the Peptidic Backbone of Glycopeptide Antibiotics

Puk¹,

Bischoff

Kittel³

et al. 2004

J Bacteriol

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The role of the putative P450 monooxygenase OxyD and the chlorination time point in the biosynthesis of the glycopeptide antibiotic balhimycin produced by Amycolatopsis balhimycina were analyzed. The oxyD gene is located directly downstream of the bhp (perhydrolase) and bpsD (nonribosomal peptide synthetase D) genes, which are involved in the synthesis of the balhimycin building block ␤-hydroxytyrosine (␤-HT). Reverse transcriptase experiments revealed that bhp, bpsD, and oxyD form an operon. oxyD was inactivated by an in-frame deletion, and the resulting mutant was unable to produce an active compound. Balhimycin production could be restored (i) by complementation with an oxyD gene, (ii) in cross-feeding studies using A. balhimycina JR1 (a null mutant with a block in the biosynthesis pathway of the building blocks hydroxy-and dihydroxyphenylglycine) as an excretor of the missing precursor, and (iii) by supplementation of ␤-HT in the growth medium. These data demonstrated an essential role of OxyD in the formation pathway of this amino acid. Liquid chromatography-electrospray ionization-mass spectrometry analysis indicated the biosynthesis of completely chlorinated balhimycin by the oxyD mutant when culture filtrates were supplemented with nonchlorinated ␤-HT. In contrast, supplementation with 3-chloro-␤-HT did not restore balhimycin production. These results indicated that the chlorination time point was later than the stage of free ␤-HT, most likely during heptapeptide synthesis.

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“…The first coupling reaction occurs between the phenol rings in residues-4 and -6 (the C-O-D ring), catalyzed by OxyB, the second aryl-ether bridge is formed between side chains of residues-2 and -4 (D-O-E ring) by OxyA, and the last biaryl coupling between the aromatic side chains of residues-5 and -7 is carried out by OxyC (AB ring, Figure-1) (Bischoff et al, 2001a;Bischoff et al, 2001b;Sussmuth et al, 1999). The crystal structures of OxyB (CYP165B1) and OxyC (CYP165C1) from the vancomycin producer A. orientalis have been reported in substrate free forms (Pylypenko et al, 2003;Zerbe et al, 2002), confirming that these proteins indeed contain a fold and heme environment typical of P450 enzymes.…”

Section: Figure-1mentioning

confidence: 99%

Chapter 19 In Vitro Studies of Phenol Coupling Enzymes Involved in Vancomycin Biosynthesis

Woithe

Geib

et al. 2009

Complex Enzymes in Microbial Natural Product Biosynthesis, Part A: Overview Articles and Peptides

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Oxidative phenol cross-linking reactions play a key role in the biosynthesis of glycopeptide antibiotics such as vancomycin. The vancomycin aglycone contains three cross-links between aromatic amino acid side-chains, which stabilize the folded backbone conformation required for binding to the target D-Ala-D-Ala dipeptide. At least the first cross-link is introduced into a peptide precursor whilst it is still bound as a thioester to a peptide carrier protein (PCP) domain (also called a thiolation domain) within the nonribosomal peptide synthetase. We described here methods for the solid-phase synthesis of peptides and their coupling to PCP domains, which may be useful for in vitro studies of cross-linking and related tailoring reactions during nonribosomal glycopeptide antibiotic biosynthesis.

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The Biosynthesis of Vancomycin-Type Glycopeptide Antibiotics-The Order of the Cyclization Steps

Cited by 137 publications

References 20 publications

Crystal Structure of OxyB, a Cytochrome P450 Implicated in an Oxidative Phenol Coupling Reaction during Vancomycin Biosynthesis

Crystal Structure of OxyB, a Cytochrome P450 Implicated in an Oxidative Phenol Coupling Reaction during Vancomycin Biosynthesis

Biosynthesis of Chloro-β-Hydroxytyrosine, a Nonproteinogenic Amino Acid of the Peptidic Backbone of Glycopeptide Antibiotics

Chapter 19 In Vitro Studies of Phenol Coupling Enzymes Involved in Vancomycin Biosynthesis

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