scholarly journals The enduracidin biosynthetic gene cluster from Streptomyces fungicidicus

Microbiology ◽  
2006 ◽  
Vol 152 (10) ◽  
pp. 2969-2983 ◽  
Author(s):  
Xihou Yin ◽  
T. Mark Zabriskie
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Gene Cluster ◽  
Genetic Locus ◽  
Biosynthetic Gene Cluster ◽  
Antibiotic Biosynthesis ◽  
Peptide Antibiotic ◽  
Amino Acid Residues ◽  
Biosynthetic Gene ◽  
Condensation Domain

The biosynthetic gene cluster for the 17 aa peptide antibiotic enduracidin has been cloned and sequenced from Streptomyces fungicidicus ATCC 21013. The 84 kb gene cluster contains 25 ORFs and is located within a 116 kb genetic locus that was fully sequenced. Targeted disruption of non-ribosomal peptide synthetase (NRPS) genes in the cluster abolished enduracidin production and confirmed function. The cluster includes four genes, endA-D, encoding two-, seven-, eight- and one-module NRPSs, respectively, and includes unique modules for the incorporation of citrulline and enduracididine. The NRPS organization generally follows the collinearity principle, and starts with a condensation domain (C domain) similar to those found in other lipopeptide systems for the coupling of an acyl group to the starting amino acid. The sixth module of EndB, corresponding to Thr8, is missing an adenylation domain (A domain) and this module is presumed to be loaded in trans by the single module protein EndD. The most striking feature of the NRPS organization is the lack of epimerization domains (E domains) in light of the fact that the product has seven d-amino acid residues. Sequence analysis reveals that C domains following modules corresponding to d-amino acids belong to a unique subset of C domains able to catalyse both epimerization and condensation reactions. Other genes directing lipid modification and activation, and formation of the non-proteinogenic amino acids 4-hydroxyphenylglycine and enduracididine are readily identified, as are genes possibly involved in regulation of antibiotic biosynthesis and export. These findings provide the basis to further genetically manipulate and improve lipodepsipeptide antibiotics via combinatorial and chemical methods.

scholarly journals Mutagenesis of NosM Leader Peptide Reveals Important Elements in Nosiheptide Biosynthesis

2016 ◽  
Vol 83 (4) ◽  
Author(s):  
Liang Jin ◽  
Xuri Wu ◽  
Yanjiu Xue ◽  
Yue Jin ◽  
Shuzhen Wang ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Gene Cluster ◽  
Genetic Manipulation ◽  
Fold Increase ◽  
Biosynthetic Gene Cluster ◽  
Leader Peptide ◽  
Amino Acid Residues ◽  
Precursor Peptide

ABSTRACT Nosiheptide, a typical member of the ribosomally synthesized and posttranslationally modified peptides (RiPPs), exhibits potent activity against multidrug-resistant Gram-positive bacterial pathogens. The precursor peptide of nosiheptide (NosM) is comprised of a leader peptide with 37 amino acids and a core peptide containing 13 amino acids. To pinpoint elements in the leader peptide that are essential for nosiheptide biosynthesis, a collection of mutants with unique sequence features, including N- and C-terminal motifs, peptide length, and specific sites in the leader peptide, was generated by mutagenesis in vivo. The effects of various mutants on nosiheptide biosynthesis were evaluated. In addition to the necessity of a conserved motif LEIS box, native length and the N-terminal 12 amino acid residues were indispensable, and single-site substitutions of these 12 amino acid residues resulted in changes ranging from a greater-than-5-fold decrease to a 2-fold increase of nosiheptide production, depending on the sites and substituted residues. Moreover, although the C-terminal motif is not conservative, significant effects of this portion on nosiheptide production were also evident. Taken together, the present results further highlight the importance of the leader peptide in nosiheptide biosynthesis, and provide new insights into the diversity and specificity of leader peptides in the biosynthesis of various RiPPs. IMPORTANCE As a representative thiopeptide, nosiheptide exhibits excellent antibacterial activity. Although the biosynthetic gene cluster and several modification steps have been revealed, the presence and roles of the leader peptide within the precursor peptide of the nosiheptide gene cluster remain elusive. Thus, identification of specific elements in the leader peptide can significantly facilitate the genetic manipulation of the gene cluster for increasing nosiheptide production or generating diverse analogues. Given the complexity of the biosynthetic process, the instability of the leader peptide, and the unavailability of intermediates, cocrystallization of intermediates, leader peptide, and modification enzymes is currently not feasible. Therefore, a mutagenesis approach was used to construct a series of leader peptide mutants to uncover a number of crucial and characteristic elements affecting nosiheptide biosynthesis, which moves a considerable distance toward a thorough understanding of the biosynthetic machinery for thiopeptides.

Daptomycin biosynthesis in Streptomyces roseosporus: cloning and analysis of the gene cluster and revision of peptide stereochemistry

Microbiology ◽  
2005 ◽  
Vol 151 (5) ◽  
pp. 1507-1523 ◽  
Author(s):  
Vivian Miao ◽  
Marie-Françoise Coëffet-LeGal ◽  
Paul Brian ◽  
Renee Brost ◽  
Julia Penn ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Gene Cluster ◽  
Binding Pocket ◽  
Biosynthetic Gene Cluster ◽  
Streptomyces Roseosporus ◽  
Cyclic Lipopeptide ◽  
Peptide Synthetase ◽  
Amino Acid Binding ◽  
Condensation Domain

Daptomycin is a 13 amino acid, cyclic lipopeptide produced by a non-ribosomal peptide synthetase (NRPS) mechanism in Streptomyces roseosporus. A 128 kb region of S. roseosporus DNA was cloned and verified by heterologous expression in Streptomyces lividans to contain the daptomycin biosynthetic gene cluster (dpt). The cloned region was completely sequenced and three genes (dptA, dptBC, dptD) encoding the three subunits of an NRPS were identified. The catalytic domains in the subunits, predicted to couple five, six or two amino acids, respectively, included a novel activation domain and amino-acid-binding pocket for incorporating the unusual amino acid l-kynurenine (Kyn), three types of condensation domains and an extra epimerase domain (E-domain) in the second module. Novel genes (dptE, dptF) whose products likely work in conjunction with a unique condensation domain to acylate the first amino acid, as well as other genes (dptI, dptJ) probably involved in supply of the non-proteinogenic amino acids l-3-methylglutamic acid and Kyn, were located next to the NRPS genes. The unexpected E-domain suggested that daptomycin would have d-Asn, rather than l-Asn, as originally assigned, and this was confirmed by comparing stereospecific synthetic peptides and the natural product both chemically and microbiologically.

scholarly journals The expansive library of jadomycins

2018 ◽  
Vol 96 (6) ◽  
pp. 495-501 ◽  
Author(s):  
Jeanna M. MacLeod ◽  
Stephanie M. Forget ◽  
David L. Jakeman
Keyword(s):  
Amino Acids ◽  
Natural Products ◽  
Amino Acid ◽  
Heterologous Expression ◽  
Gene Cluster ◽  
Gene Deletion ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Synthetic Derivatives ◽  
Subsequent Identification

The jadomycin family of natural products was discovered from Streptomyces venezuelae ISP5230 in the 1990s. Subsequent identification of the biosynthetic gene cluster along with synthetic efforts established that incorporation of an amino acid into the polyaromatic angucycline core occurs non-enzymatically. Over two decades, the precursor-directed biosynthetic potential of the jadomycins has been heavily exploited, generating a library exceeding 70 compounds. This review compiles the jadomycins that have been isolated and characterized to date; these include jadomycins incorporating proteinogenic and non-proteinogenic amino acids, semi-synthetic derivatives, biosynthetic shunt products, compounds isolated in structural gene deletion studies, and deoxysugar sugar variant jadomycins produced by deletion or heterologous expression of sugar biosynthetic genes.

scholarly journals Pentaminomycins F and G, First Non-Ribosomal Peptides Containing 2-Pyridylalanine

2020 ◽  
Author(s):  
Daniel Carretero Molina ◽  
Francisco Javier Ortiz-Lopez ◽  
Jesús Martín ◽  
Ignacio González ◽  
Marina Sánchez-Hidalgo ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Amino Acid ◽  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Peptide Synthetase ◽  
Non Ribosomal Peptides

Pentaminomycins F-H, a group of three new hydroxyarginine-containing cyclic pentapeptides, were isolated from cultures of a <i>Streptomyces cacaoi</i> subsp. <i>cacaoi</i> strain along with the known pentaminomycins A-E. The structures of the new peptides were determined by a combination of mass spectrometry and NMR and Marfey's analyses. Among them, pentaminomycins F and G were shown to contain in their structures the rare amino acid 3-(2-pyridyl)-alanine. This finding represents the first reported example of non-ribosomal peptides containing this residue. The LDLLD chiral sequence found for the three compounds was in agreement with that reported for previously isolated pentaminomycins and consistent with the epimerization domains present in the putative non-robosomal peptide synthetase (NRPS) biosynthetic gene cluster.<br>

scholarly journals Biosynthetic Investigations of Lactonamycin and Lactonamycin Z: Cloning of the Biosynthetic Gene Clusters and Discovery of an Unusual Starter Unit

2007 ◽  
Vol 52 (2) ◽  
pp. 574-585 ◽  
Author(s):  
Xiujun Zhang ◽  
Lawrence B. Alemany ◽  
Hans-Peter Fiedler ◽  
Michael Goodfellow ◽  
Ronald J. Parry
Keyword(s):  
Gene Cluster ◽  
Genetic Locus ◽  
Gene Clusters ◽  
Biosynthetic Gene Cluster ◽  
Open Reading Frames ◽  
Antibacterial Drug ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Starter Unit ◽  
High Degree

ABSTRACT The antibiotics lactonamycin and lactonamycin Z provide attractive leads for antibacterial drug development. Both antibiotics contain a novel aglycone core called lactonamycinone. To gain insight into lactonamycinone biosynthesis, cloning and precursor incorporation experiments were undertaken. The lactonamycin gene cluster was initially cloned from Streptomyces rishiriensis. Sequencing of ca. 61 kb of S. rishiriensis DNA revealed the presence of 57 open reading frames. These included genes coding for the biosynthesis of l-rhodinose, the sugar found in lactonamycin, and genes similar to those in the tetracenomycin biosynthetic gene cluster. Since lactonamycin production by S. rishiriensis could not be sustained, additional proof for the identity of the S. rishiriensis cluster was obtained by cloning the lactonamycin Z gene cluster from Streptomyces sanglieri. Partial sequencing of the S. sanglieri cluster revealed 15 genes that exhibited a very high degree of similarity to genes within the lactonamycin cluster, as well as an identical organization. Double-crossover disruption of one gene in the S. sanglieri cluster abolished lactonamycin Z production, and production was restored by complementation. These results confirm the identity of the genetic locus cloned from S. sanglieri and indicate that the highly similar locus in S. rishiriensis encodes lactonamycin biosynthetic genes. Precursor incorporation experiments with S. sanglieri revealed that lactonamycinone is biosynthesized in an unusual manner whereby glycine or a glycine derivative serves as a starter unit that is extended by nine acetate units. Analysis of the gene clusters and of the precursor incorporation data suggested a hypothetical scheme for lactonamycinone biosynthesis.

scholarly journals Discovery of Gene Cluster for Mycosporine-Like Amino Acid Biosynthesis from Actinomycetales Microorganisms and Production of a Novel Mycosporine-Like Amino Acid by Heterologous Expression

2014 ◽  
Vol 80 (16) ◽  
pp. 5028-5036 ◽  
Author(s):  
Kiyoko T. Miyamoto ◽  
Mamoru Komatsu ◽  
Haruo Ikeda
Keyword(s):  
Amino Acid ◽  
Gene Cluster ◽  
Gene Clusters ◽  
Biosynthetic Gene Cluster ◽  
Dependent Manner ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Gram Positive ◽  
Content Type ◽  
Gram Positive Bacteria

ABSTRACTMycosporines and mycosporine-like amino acids (MAAs), including shinorine (mycosporine-glycine-serine) and porphyra-334 (mycosporine-glycine-threonine), are UV-absorbing compounds produced by cyanobacteria, fungi, and marine micro- and macroalgae. These MAAs have the ability to protect these organisms from damage by environmental UV radiation. Although no reports have described the production of MAAs and the corresponding genes involved in MAA biosynthesis from Gram-positive bacteria to date, genome mining of the Gram-positive bacterial database revealed that two microorganisms belonging to the orderActinomycetales,Actinosynnema mirumDSM 43827 andPseudonocardiasp. strain P1, possess a gene cluster homologous to the biosynthetic gene clusters identified from cyanobacteria. When the two strains were grown in liquid culture,Pseudonocardiasp. accumulated a very small amount of MAA-like compound in a medium-dependent manner, whereasA. mirumdid not produce MAAs under any culture conditions, indicating that the biosynthetic gene cluster ofA. mirumwas in a cryptic state in this microorganism. In order to characterize these biosynthetic gene clusters, each biosynthetic gene cluster was heterologously expressed in an engineered host,Streptomyces avermitilisSUKA22. Since the resultant transformants carrying the entire biosynthetic gene cluster controlled by an alternative promoter produced mainly shinorine, this is the first confirmation of a biosynthetic gene cluster for MAA from Gram-positive bacteria. Furthermore,S. avermitilisSUKA22 transformants carrying the biosynthetic gene cluster for MAA ofA. mirumaccumulated not only shinorine and porphyra-334 but also a novel MAA. Structure elucidation revealed that the novel MAA is mycosporine-glycine-alanine, which substitutesl-alanine for thel-serine of shinorine.

scholarly journals In Vivo Analysis of the Regulatory Genes in the Nystatin Biosynthetic Gene Cluster of Streptomyces noursei ATCC 11455 Reveals Their Differential Control Over Antibiotic Biosynthesis

2004 ◽  
Vol 186 (5) ◽  
pp. 1345-1354 ◽  
Author(s):  
Olga N. Sekurova ◽  
Trygve Brautaset ◽  
Håvard Sletta ◽  
Sven E. F. Borgos ◽  
Øyvind M. Jakobsen ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Regulatory Genes ◽  
Antibiotic Biosynthesis ◽  
Biosynthetic Gene ◽  
Promoter Regions ◽  
Streptomyces Noursei ◽  
In Vivo Analysis ◽  
Differential Control

ABSTRACT Six putative regulatory genes are located at the flank of the nystatin biosynthetic gene cluster in Streptomyces noursei ATCC 11455. Gene inactivation and complementation experiments revealed that nysRI, nysRII, nysRIII, and nysRIV are necessary for efficient nystatin production, whereas no significant roles could be demonstrated for the other two regulatory genes. To determine the in vivo targets for the NysR regulators, chromosomal integration vectors with the xylE reporter gene under the control of seven putative promoter regions upstream of the nystatin structural and regulatory genes were constructed. Expression analyses of the resulting vectors in the S. noursei wild-type strain and regulatory mutants revealed that the four regulators differentially affect certain promoters. According to these analyses, genes responsible for initiation of nystatin biosynthesis and antibiotic transport were the major targets for regulation. Data from cross-complementation experiments showed that nysR genes could in some cases substitute for each other, suggesting a functional hierarchy of the regulators and implying a cascade-like mechanism of regulation of nystatin biosynthesis.

scholarly journals Biosynthetic Pathway for Mannopeptimycins, Lipoglycopeptide Antibiotics Active against Drug-Resistant Gram-Positive Pathogens

2006 ◽  
Vol 50 (6) ◽  
pp. 2167-2177 ◽  
Author(s):  
Nathan A. Magarvey ◽  
Brad Haltli ◽  
Min He ◽  
Michael Greenstein ◽  
John A. Hucul
Keyword(s):  
Amino Acid ◽  
Gene Cluster ◽  
Biosynthetic Pathway ◽  
Domain Analysis ◽  
Biosynthetic Gene Cluster ◽  
Multidrug Resistant ◽  
Drug Resistant ◽  
Biosynthetic Gene ◽  
Peptide Synthetases ◽  
Multidrug Resistant Pathogens

ABSTRACT The mannopeptimycins are a novel class of lipoglycopeptide antibiotics active against multidrug-resistant pathogens with potential as clinically useful antibacterials. This report is the first to describe the biosynthesis of this novel class of mannosylated lipoglycopeptides. Included here are the cloning, sequencing, annotation, and manipulation of the mannopeptimycin biosynthetic gene cluster from Streptomyces hygroscopicus NRRL 30439. Encoded by genes within the mannopeptimycin biosynthetic gene cluster are enzymes responsible for the generation of the hexapeptide core (nonribosomal peptide synthetases [NRPS]) and tailoring reactions (mannosylation, isovalerylation, hydroxylation, and methylation). The NRPS system is noncanonical in that it has six modules utilizing only five amino acid-specific adenylation domains and it lacks a prototypical NRPS macrocyclizing thioesterase domain. Analysis of the mannopeptimycin gene cluster and its engineering has elucidated the mannopeptimycin biosynthetic pathway and provides the framework to make new and improved mannopeptimycins biosynthetically.

scholarly journals Biosynthesis of cittilins, unusual ribosomally synthesized and post-translationally modified peptides from Myxococcus xanthus

2020 ◽  
Author(s):  
Joachim J. Hug ◽  
Jan Dastbaz ◽  
Sebastian Adam ◽  
Ole Revermann ◽  
Jesko Koehnke ◽  
...  
Keyword(s):  
Amino Acid ◽  
Gene Cluster ◽  
Biochemical Characterization ◽  
Biosynthetic Gene Cluster ◽  
Cytochrome P450 Enzyme ◽  
Biosynthetic Gene ◽  
Aryl Ether ◽  
Precursor Peptide ◽  
Carbon Storage Regulator

AbstractCittilins are secondary metabolites from myxobacteria comprised of three L-tyrosines and one L-isoleucine forming a bicyclic tetrapeptide scaffold with biaryl and aryl-oxygen-aryl ether bonds. Here we reveal that cittilins belong to the ribosomally synthesized and post-translationally modified peptide (RiPP) family of natural products, for which only the crocagins have been reported from myxobacteria. A 27 amino acid precursor peptide harbors a C-terminal four amino acid core peptide, which is enzymatically modified and finally exported to yield cittilins. The small biosynthetic gene cluster responsible for cittilin biosynthesis also encodes a cytochrome P450 enzyme and a methyltransferase, whereas a gene encoding a prolyl endopeptidase for the cleavage of the precursor peptide is located outside of the cittilin biosynthetic gene cluster. We confirm the roles of the biosynthetic genes responsible for the formation of cittilins using targeted gene inactivation and heterologous expression in Streptomyces. We also report first steps towards the biochemical characterization of the proposed biosynthetic pathway in vitro. An investigation of the cellular uptake properties of cittilin A connected it to a potential biological function as an inhibitor of the prokaryotic carbon storage regulator A (CsrA).Abstract Figure

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