Analysis of the nourseothricin-resistance gene (nat) of Streptomyces noursei

Krügel, H.; Fiedler, G.; Haupt, Ina; Sarfert, Eva; Simon, H

doi:10.1016/0378-1119(88)90559-8

Cited by 37 publications

(29 citation statements)

References 17 publications

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“…The 162 bp fragment from pBR322 was that described previously [16]. The 169 bp fragment containing a high GC content was prepared from the recombinant plasmid pUC19 carrying the nourseothricyl-acetyl-transferase (nat) gene of S. noursei inserted into the BamHI site [17]. The fragment was derived by digestion of the plasmid with PstI-KpnI and subsequent treatment of the 250 bp fragment with SalI.…”

Section: Dnamentioning

confidence: 99%

Biochemical complementation studies in vitro of gyrase subunits from different species

1995

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To investigate the functional equivalence of DNA gyrase subunits from different bacterial sources hybrid enzymes were formed using purified A and B subunits from three species of Streptomycetes, E. coli and B. subtilis. The activity of gyrase hybrids composed of heterologous gyr A and gyr B proteins and of the gyrases containing homologous subunits was characterized by binding studies and a cleavage assay with two different DNA fragments. Likewise the enzyme activity was monitored by the super-coiling and relaxation assay with pBR322 DNA. We found that cleavage reactions are largely determined by the source of the gyr B subunits whereas DNA supercoiling and relaxation reactions of pBR322 catalyzed by DNA gyrase are limited to a combination of homologous A and B subunits or of heterologous A and B subunits from the taxonomically related bacteria Streptomycetes.

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Section: Dnamentioning

confidence: 99%

Biochemical complementation studies in vitro of gyrase subunits from different species

1995

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“…2) (20). SATs in different ST-producing strains are well known to catalyze the acetyl-CoA-dependent acetylation at the ␤-amino group of the ␤-Lys moiety in ST-F, conferring ST resistance (31,32). Although no reports about the substrate specificity of SATs have been published, the orf1 gene product (Orf1-SAT) was considered to be a candidate for the enzyme that acetylates glycylthricin.…”

Section: Identification Of the Bd-12-biosynthetic Gene Cluster From Smentioning

confidence: 99%

tRNA-Dependent Aminoacylation of an Amino Sugar Intermediate in the Biosynthesis of a Streptothricin-Related Antibiotic

Maruyama

Niikura

Izumikawa

et al. 2016

Appl Environ Microbiol

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The antibiotic streptothricin (ST) possesses an amino sugar bound to an L-␤-lysine (␤-Lys) residue via a peptide bond. The peptide bond formation has been shown to be catalyzed by a nonribosomal peptide synthetase (NRPS) during ST biosynthesis. The focus of this study is the closely related ST analogue BD-12, which carries a glycine-derived side chain rather than a ␤-Lys residue. Here, in Streptomyces luteocolor NBRC13826, we describe our biosynthetic studies of BD-12, which revealed that the peptide bond between the amino sugar and the glycine residue is catalyzed by a Fem-like enzyme (Orf11) in a tRNA-dependent manner rather than by an NRPS. Although there have been several reports of peptide bond-forming tRNA-dependent enzymes, to our knowledge, Orf11 is the first enzyme that can accept an amino sugar as a substrate. Our findings clearly demonstrate that the structural diversity of the side chains of ST-type compounds in nature is generated in an unusual manner via two distinct peptide bond-forming mechanisms. Moreover, the identification and functional analysis of Orf11 resulted in not only the production of new ST-related compounds, but also the provision of new insights into the structure-activity relationship of the ST-related antibiotics. (1), STs with an oligo(␤-Lys) consisting of two to seven residues have been identified (Fig. 1). ST-F inhibits protein biosynthesis in prokaryotic cells (2), and STs carrying the longer oligo(␤-Lys) side chains show higher levels of antibacterial activity. Moreover, STs strongly inhibit the growth of eukaryotes, such as yeasts (3-5), fungi (6), protozoa (7), insects (8), plants (9), and mammals (10-13). Although STs have been used effectively as selective agents for recombinant DNA work in some of these organisms, STs are not currently used therapeutically due to their inherent toxicity. In addition to the STs, it has been reported that Streptomyces strains produce ST analogues that possess a glycinederived side chain rather than the ␤-Lys residue: 15), citromycin (16,17), glycinothricin (18), A-269A (19), and A-269A= (19) (Fig. 1). These analogues display potent antibacterial activities, although their molecular targets remain unclear. Also, like the STs, the ST analogues are not used clinically due to their toxicity.We previously identified the ST-biosynthetic gene cluster (accession no. AB684619) in Streptomyces rochei NBRC 12908 (Fig. 2) and elucidated the biosynthetic mechanisms of the oligo(␤-Lys) side chains (20). Nonribosomal peptide synthetases (NRPSs) are known to catalyze the assembly of a myriad of structurally complex peptide natural products (21). However, in the previous study, we identified three unique stand-alone NRPSs among the ST-biosynthetic enzymes and showed that they assembled the structurally simple peptide oligo(␤-Lys) (Fig. 3A). The biosynthesis is initiated by adenylation of ␤-Lys in Orf5 (stand-alone adenylation [A] domain), and the resulting L-␤-lysyl-O-AMP is loaded

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“…Strains-Based on the highly conserved amino acid sequence of the NATs of S. lavendulae (16), S. rochei (17), and S. noursei (18,19), we designed primers 5Ј-…”

Section: Pcr Amplification Of the Genes Encoding Nats In The Streptommentioning

confidence: 99%

“…1). In fact, in ST producers such as S. lavendulae (16), Streptomyces rochei (17), and Streptomyces noursei (18,19), the ST-resistance genes encoding N-acetyltransferase (NAT) have been identified and their role in selfresistance against their own STs has been investigated. Based on this resistance mechanism and the fact that streptothricin D (ST-D) is a more effective antibiotic than streptothricin F (ST-F), the moiety of ␤-lysine(s) has been shown to play a crucial role in antibiotic activity.…”

mentioning

confidence: 99%

A Novel Enzyme Conferring Streptothricin Resistance Alters the Toxicity of Streptothricin D from Broad-spectrum to Bacteria-specific

Hamano

Matsuura

Kitamura

et al. 2006

Journal of Biological Chemistry

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Analysis of the nourseothricin-resistance gene (nat) of Streptomyces noursei

Cited by 37 publications

References 17 publications

Biochemical complementation studies in vitro of gyrase subunits from different species

Biochemical complementation studies in vitro of gyrase subunits from different species

tRNA-Dependent Aminoacylation of an Amino Sugar Intermediate in the Biosynthesis of a Streptothricin-Related Antibiotic

A Novel Enzyme Conferring Streptothricin Resistance Alters the Toxicity of Streptothricin D from Broad-spectrum to Bacteria-specific

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