The Biosynthetic Gene Cluster for a Monocyclic β-Lactam Antibiotic, Nocardicin A

Gunsior, Michele; Breazeale, Steven D.; Lind, Amanda J; Ravel, Jacques; Janc, James W.; Townsend, Craig A.

doi:10.1016/j.chembiol.2004.04.012

Cited by 68 publications

(110 citation statements)

References 62 publications

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“…A UV-visible spectrum in ddH 2 Complementation of nocJ::apra N. uniformis-nocJ was amplified by PCR from cosmid DNA (17). The forward primer (5Ј-AACATAT-GGGCGCGCTGCCGCGCGTG-3Ј) possessed an NdeI restriction site (underlined) and the native GTG start codon was replaced with ATG.…”

Section: Methodsmentioning

confidence: 99%

“…Construction of nocJ::apra N. uniformis-The gene nocJ was amplified from cosmid DNA by PCR (17). The forward primer (5Ј-AAGGATC-CGTGGGCGCGCTG-3Ј) possessed a BamHI restriction site (underlined) and contained the wild-type start codon.…”

Section: Methodsmentioning

confidence: 99%

“…Vectors pIJ4070 and pULVK2 were generous gifts of Professors Mervyn Bibb (John Innes Center, Norwich, England) and Juan F. Martín (Area of Microbiology, University of León, León, Spain), respectively. Vector pULVK2T was described elsewhere (17). Wild-type N. uniformis subsp.…”

Section: Methodsmentioning

confidence: 99%

“…Notably present in the cluster are two non-ribosomal peptide synthetases (NRPSs), genes required for generation of the non-proteinogenic amino acid p-(hydroxyphenyl)glycine, (17) nocL, which encodes a cytochrome P450 responsible for oxime formation in the nocardicins, (18) and nat providing the side chain AdoMet-dependent transferase. Also present in the cluster is an open reading frame, nocJ, whose predicted amino acid sequence shows similarity to the pyridoxal 5Ј-phosphate (PLP) 1 -dependent 1-aminocyclopropane-1-carboxylate (ACC) deaminases (17).…”

mentioning

confidence: 99%

“…Reverse genetics techniques from the AdoMet-dependent 3-amino-3-carboxypropyl transferase (Nat) have allowed the nocardicin biosynthetic gene cluster to be identified and characterized from N. uniformis (16,17). Notably present in the cluster are two non-ribosomal peptide synthetases (NRPSs), genes required for generation of the non-proteinogenic amino acid p-(hydroxyphenyl)glycine, (17) nocL, which encodes a cytochrome P450 responsible for oxime formation in the nocardicins, (18) and nat providing the side chain AdoMet-dependent transferase.…”

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confidence: 99%

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Mutational Analysis and Characterization of Nocardicin C-9′ Epimerase

Kelly

Townsend

2004

Journal of Biological Chemistry

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The biosynthetic gene cluster for the nocardicin A producer Nocardia uniformis subsp. tsuyamanensis ATCC 21806 was recently identified. Nocardicin A is the most potent of a series of monocyclic ␤-lactam antibiotics produced by this organism. Its activity has been attributed to a syn-configured oxime moiety and a D-homoseryl side chain attached through an unusual ether linkage to the core nocardicin framework. Notably present in the nocardicin biosynthetic gene cluster is nocJ, encoding a protein with sequence similarity to the pyridoxal 5 -phosphate (PLP)-dependent 1-aminocyclopropane-1-carboxylic acid deaminases. Insertional mutagenesis of nocJ abolished nocardicin A production, while the L-homoseryl isomer, isonocardicin A, was still observed. Expression of the disrupted nocJ gene in trans was sufficient to restore production of nocardicin A in the disruption mutant. Heterologous expression, purification, and in vitro characterization of NocJ by UV spectroscopy, cofactor reduction, chiral HPLC analysis of the products and their exchange behavior in deuterium oxide led to confirmation of its role as the PLP-dependent nocardicin C-9 epimerase responsible for interconversion of the nocardicin homoseryl side chain in both nocardicin A with isonocardicin A, and nocardicin C with isonocardicin C. NocJ is the first member of a new class of ␤-lactam aminoacyl side chain epimerases, the first two classes being the evolutionarily distinct prokaryotic PLP-dependent isopenicillin N epimerase and the fungal isopenicillin N epimerase two protein system.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Mutational Analysis and Characterization of Nocardicin C-9′ Epimerase

Kelly

Townsend

2004

Journal of Biological Chemistry

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β‐Lactam Antibiotics

Testero

Fisher

Mobashery

2010

Burger's Medicinal Chemistry and Drug Discovery

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The β‐lactam classes of antibacterials are preeminent in the treatment of bacterial infection due to their unparalleled clinical efficacy and clinical safety. Following the discovery of the penicillins, successive β‐lactam drug discovery has added the cephalosporin, penem cephamycin, clavulanate, monobactam, nocardicin, and carbapenem subclasses. The driving force behind much of this era of discovery is the staggering ability of pathogenic bacteria to adapt previous generations of the β‐lactam by the acquisition and expression of resistance mechanisms. Although many factors contribute to β‐lactam resistance, alterations to the molecular targets of the β‐lactams (the penicillin binding proteins) and the use of enzymes (the β‐lactamases) capable of the hydrolytic deactivation of the β‐lactams are paramount. This review traces the historical development of β‐lactam drug discovery, with emphasis on the most recent progress in the medicinal chemistry, biochemistry, and microbiology of the β‐lactams leading to the discovery of new generation β‐lactam antibacterials effective against the Gram‐negative and ‐positive bacterial pathogens of current medical concern.

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Four‐Membered Heterocyclic Rings and Their Fused Derivatives

2015

Biosynthesis of Heterocycles

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The Biosynthetic Gene Cluster for a Monocyclic β-Lactam Antibiotic, Nocardicin A

Cited by 68 publications

References 62 publications

Mutational Analysis and Characterization of Nocardicin C-9′ Epimerase

Mutational Analysis and Characterization of Nocardicin C-9′ Epimerase

β‐Lactam Antibiotics

Four‐Membered Heterocyclic Rings and Their Fused Derivatives

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