scholarly journals Nitrate reductase from the magnetotactic bacteriumMagnetospirillum magnetotacticumMS-1: purification and sequence analyses

2003 ◽  
Vol 49 (3) ◽  
pp. 197-206 ◽  
Author(s):  
Azuma Taoka ◽  
Katsuhiko Yoshimatsu ◽  
Masaaki Kanemori ◽  
Yoshihiro Fukumori
Keyword(s):  
Nitrate Reductase ◽  
Gene Cluster ◽  
Phylogenetic Analyses ◽  
Regulatory Protein ◽  
Heme Iron ◽  
Open Reading Frames ◽  
Non Heme Iron ◽  
Close Relationship ◽  
Paracoccus Pantotrophus ◽  
Magnetospirillum Magnetotacticum

We purified the nitrate reductase from the soluble fraction of Magnetospirillum magnetotacticum MS-1. The enzyme was composed of 86- and 17-kDa subunits and contained molybdenum, non-heme iron, and heme c. These properties are very similar to those of the periplasmic nitrate reductase found in Paracoccus pantotrophus. The M. magnetotacticum nap locus was clustered in seven open reading frames, napFDAGHBC. The phylogenetic analyses of NapA, NapB, and NapC suggested a close relationship between M. magnetotacticum nap genes and Escherichia coli nap genes, which is not consistent with the 16S rDNA data. This is the first finding that the α subclass of Proteobacteria possesses a napFDAGHBC-type nap gene cluster. The nap gene cluster had putative fumarate and nitrate reduction regulatory protein (Fnr) and NarL protein binding sites. Furthermore, we investigated the effect of molybdate deficiency in medium on the total iron content of the magnetosome fraction and discussed the physiological function of nitrate reductase in relation to the magnetite synthesis in M. magnetotacticum.Key words: nitrate reductase, magnetotactic bacteria, denitrification, horizontal gene transfer.

scholarly journals Characterization of the DNA polymerase loci of the novel porcine lymphotropic herpesviruses 1 and 2 in domestic and feral pigs

1999 ◽  
Vol 80 (12) ◽  
pp. 3199-3205 ◽  
Author(s):  
Sven Ulrich ◽  
Michael Goltz ◽  
Bernhard Ehlers
Keyword(s):  
Dna Polymerase ◽  
Phylogenetic Analyses ◽  
Open Reading Frames ◽  
The Novel ◽  
Feral Pigs ◽  
Virus Transfer ◽  
Close Relationship ◽  
Flanking Sequences ◽  
Cpg Dinucleotide

Two novel porcine gammaherpesviruses, porcine lymphotropic herpesviruses 1 and 2 (PLHV-1 and -2), have been detected by amplification of short DNA polymerase (DPOL) sequences from blood and spleen of domestic pigs while searching for unknown herpesviruses in pigs as possible risk factors in xenotransplantation. In the present study, the DPOL genes of the two viruses and the open reading frames (ORFs) that follow in the downstream direction were amplified by PCR-based genome walking from adaptor-ligated restriction fragment libraries of porcine spleen samples. The sequences determined for the two PLHVs exhibited a very low G+C content (37 mol%) and a marked suppression of the CpG dinucleotide frequency. The DPOL proteins encoded were 95% identical and showed a close relationship (60% identity) to the DPOL protein of a ruminant gammaherpesvirus, alcelaphine herpesvirus 1 (AlHV-1). This was confirmed by phylogenetic analyses of the conserved regions of the two PLHV DPOL proteins. The PLHV ORFs downstream of DPOL exhibited 83% identity to each other and ≫50% similarity to ORF A5, the position equivalent of AlHV-1. From these data, the PLHVs can be firmly classified to the subfamily Gammaherpesvirinae. To find a natural reservoir for the PLHVs, organs of feral pigs were screened with five different PCR assays, targetting either the DPOL gene or 3′-flanking sequences. In all samples, PLHV sequences were detected that originated predominantly from PLHV-2, suggesting the possibility of virus transfer between feral and domestic pig populations.

scholarly journals Evolutionary Relationship between Chlorocatechol Catabolic Enzymes from Rhodococcus opacus 1CP and Their Counterparts in Proteobacteria: Sequence Divergence and Functional Convergence

1998 ◽  
Vol 180 (5) ◽  
pp. 1082-1094 ◽  
Author(s):  
Dirk Eulberg ◽  
Elena M. Kourbatova ◽  
Ludmila A. Golovleva ◽  
Michael Schlömann
Keyword(s):  
Gene Cluster ◽  
Regulatory Protein ◽  
Evolutionary Relationship ◽  
Sequence Divergence ◽  
Functional Adaptation ◽  
Open Reading Frames ◽  
Rhodococcus Opacus ◽  
Sequence Comparisons ◽  
Catabolic Enzymes ◽  
Chloromuconate Cycloisomerase

Biochemical investigations of the muconate and chloromuconate cycloisomerases from the chlorophenol-utilizing strainRhodococcus opacus (erythropolis) 1CP had previously indicated that the chlorocatechol catabolic pathway of this strain may have developed independently from the corresponding pathways of proteobacteria. To test this hypothesis, we cloned the chlorocatechol catabolic gene cluster of strain 1CP by using PCR with primers derived from sequences of N termini and peptides of purified chlorocatechol 1,2-dioxygenase and chloromuconate cycloisomerase. Sequencing of the clones revealed that they comprise different parts of the same gene cluster in which five open reading frames have been identified. The clcB gene for chloromuconate cycloisomerase is transcribed divergently from a gene which codes for a LysR-type regulatory protein, the presumed ClcR. Downstream of clcRbut separated from it by 222 bp, we detected the clcA andclcD genes, which could unambiguously be assigned to chlorocatechol 1,2-dioxygenase and dienelactone hydrolase. A gene coding for a maleylacetate reductase could not be detected. Instead, the product encoded by the fifth open reading frame turned out to be homologous to transposition-related proteins of IS1031 and Tn4811. Sequence comparisons of ClcA and ClcB to other 1,2-dioxygenases and cycloisomerases, respectively, clearly showed that the chlorocatechol catabolic enzymes of R. opacus 1CP represent different branches in the dendrograms than their proteobacterial counterparts. Thus, while the sequences diverged, the functional adaptation to efficient chlorocatechol metabolization occurred independently in proteobacteria and gram-positive bacteria, that is, by functionally convergent evolution.

scholarly journals Nostophycin Biosynthesis Is Directed by a Hybrid Polyketide Synthase-Nonribosomal Peptide Synthetase in the Toxic Cyanobacterium Nostoc sp. Strain 152

2011 ◽  
Vol 77 (22) ◽  
pp. 8034-8040 ◽  
Author(s):  
David P. Fewer ◽  
Julia Österholm ◽  
Leo Rouhiainen ◽  
Jouni Jokela ◽  
Matti Wahlsten ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Polyketide Synthase ◽  
Catalytic Domain ◽  
Phylogenetic Analyses ◽  
Biosynthetic Gene Cluster ◽  
Nonribosomal Peptide Synthetase ◽  
Open Reading Frames ◽  
Nonribosomal Peptide ◽  
Peptide Synthetase ◽  
Biochemical Analyses

ABSTRACTCyanobacteria are a rich source of natural products with interesting pharmaceutical properties. Here, we report the identification, sequencing, annotation, and biochemical analysis of the nostophycin (npn) biosynthetic gene cluster. Thenpngene cluster spans 45.1 kb and consists of three open reading frames encoding a polyketide synthase, a mixed polyketide nonribosomal peptide synthetase, and a nonribosomal peptide synthetase. The genetic architecture and catalytic domain organization of the proteins are colinear in arrangement, with the putative order of the biosynthetic assembly of the cyclic heptapeptide. NpnB contains an embedded monooxygenase domain linking nonribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) catalytic domains and predicted here to hydroxylate the nostophycin during assembly. Expression of the adenylation domains and subsequent substrate specificity assays support the involvement of this cluster in nostophycin biosynthesis. Biochemical analyses suggest that the loading substrate of NpnA is likely to be a phenylpropanoic acid necessitating deletion of a carbon atom to explain the biosynthesis of nostophycin. Biosyntheses of nostophycin and microcystin resemble each other, but the phylogenetic analyses suggest that they are distantly related to one another.

Nitrate-reductase electron-transport cofactors in Veillonella alcalescens

1977 ◽  
Vol 23 (11) ◽  
pp. 1562-1567 ◽  
Author(s):  
K. L. Ruoff ◽  
E. A. Delwiche
Keyword(s):  
Electron Transport ◽  
Nitrate Reductase ◽  
Growth Medium ◽  
Doubling Time ◽  
Growth Yield ◽  
Heme Iron ◽  
Deficient Mutant ◽  
Non Heme Iron ◽  
Reducing Activity

Studies on the effects of inhibitors of the nitrate-reducing activity of Veillonella alcalescens extracts suggest the participation of a naphthoquinone, a b-type cytochrome, and non-heme iron in electron transport to nitrate. A nitrate-reductase-deficient mutant displayed a longer doubling time and a decreased molar growth yield on nitrate media. This mutant was phenotypically restored by the addition of molybdate to the growth medium, giving evidence for the functioning of molybdenum in the nitrate-reductase enzyme of V. alcalescens.

scholarly journals Occurrence of a Strain of Texas pepper virus in Tabasco and Habanero Pepper in Costa Rica

Plant Disease ◽  
2000 ◽  
Vol 84 (2) ◽  
pp. 168-172 ◽  
Author(s):  
Pongtharin Lotrakul ◽  
Rodrigo A. Valverde ◽  
Rodolfo De La Torre ◽  
Jeonggu Sim ◽  
Alvaro Gomez
Keyword(s):  
Costa Rica ◽  
Phylogenetic Analyses ◽  
Viral Disease ◽  
Complete Sequence ◽  
Open Reading Frames ◽  
Degenerate Primers ◽  
Habanero Pepper ◽  
Close Relationship ◽  
Viruslike Particles ◽  
Pepper Plants

A viral disease causing severe leaf malformation and yellow mottle on Tabasco (Capsicum frutescens) and Habanero (C. chinense) pepper plants was observed in 1997 on farms in southwestern Costa Rica. Whiteflies (Bemisia tabaci) were present on affected farms and transmitted the putative virus. Total DNA was extracted from a whitefly-transmitted isolate, and polymerase chain reaction (PCR) was performed using degenerate primers. The expected PCR product (550 bp) was obtained, suggesting the presence of a geminivirus. This was confirmed by Southern analysis using a geminivirus-specific probe. The virus was mechanically transmitted from pepper to pepper. Electron microscopy of ultrathin sections from infected Tabasco pepper plants revealed fibrillar rings and viruslike particles in the nucleus of the vascular parenchyma cells. The sequence of DNA A was obtained from three overlapping PCR fragments amplified using three pairs of degenerate primers; PAL1v1978/PAR1c496, PCRc1/AV494, and PCRv181/ AC1048. The complete sequence of DNA A of this begomovirus consisted of 2,619 bp (GenBank accession number: AF149227) containing four open reading frames (ORF). The nucleotide sequence of the virus was 92.3% identical to DNA A of the Tamaulipas strain of Texas pepper virus (TPV-TAM). Phylogenetic analyses using AC1 and AV1 nucleotide sequences also indicated a close relationship between this virus and TPV. Based on the biological characteristics, the high percentage of nucleotide and derived amino acid sequence identities, and phyloge-netic analyses, we concluded that this virus is a distinct strain of TPV, and designated it as the Costa Rica strain. This is the first report of TPV in Costa Rica.

scholarly journals Analysis of the Nitric Oxide-sensing Non-heme Iron Center in the NorR Regulatory Protein

2007 ◽  
Vol 283 (2) ◽  
pp. 908-918 ◽  
Author(s):  
Nicholas P. Tucker ◽  
Benoît D'Autréaux ◽  
Faridoon K. Yousafzai ◽  
Shirley A. Fairhurst ◽  
Stephen Spiro ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Regulatory Protein ◽  
Heme Iron ◽  
Non Heme Iron ◽  
Iron Center

scholarly journals Genomic characterization of the first insectivoran papillomavirus reveals an unusually long, second non-coding region and indicates a close relationship to Betapapillomavirus

2009 ◽  
Vol 90 (3) ◽  
pp. 626-633 ◽  
Author(s):  
Eric Schulz ◽  
Marc Gottschling ◽  
Ignacio G. Bravo ◽  
Ullrich Wittstatt ◽  
Eggert Stockfleth ◽  
...  
Keyword(s):  
Biological Diversity ◽  
Sequence Data ◽  
Phylogenetic Analyses ◽  
Open Reading Frames ◽  
Follicle Cells ◽  
Modular Organization ◽  
Coding Region ◽  
Close Relationship ◽  
European Hedgehog ◽  
History Of

Knowledge about biological diversity is the prerequisite to reliably reconstruct the evolution of pathogens such as papillomaviruses (PV). However, complete genomes of non-human PV have only been cloned and sequenced from 8 out of 18 orders within the Placentalia, although the host-specific variety of PV is considered much larger. We isolated and sequenced the complete genome of the first insectivoran PV type from hair follicle cells of the European hedgehog (Erinaceus europaeus), designated EHPV. We conducted phylogenetic analyses (maximum-likelihood criterion and Bayesian inference) with the genomic information of a systematically representative set of 67 PV types including EHPV. As inferred from amino acid sequence data of the separate genes E1, E2 and L1 as well as of the gene combination E6–E7–E1–E2–L1, EHPV clustered within the β-γ-π-Ξ-PV supertaxon and constituted the closest relative of genus Betapapillomavirus infecting primates. Beside the typical organization of the PV genome, EHPV exhibited a 1172 bp, non-coding region between the E2 and the L2 open reading frames. This trait has been previously described for the only distantly related Lambdapapillomavirus, but a common evolutionary origin of both non-coding regions is unlikely. Our results underscore the modular organization of the PV genome and the complex natural history of PV.

scholarly journals The Mithramycin Gene Cluster of Streptomyces argillaceus Contains a Positive Regulatory Gene and Two Repeated DNA Sequences That Are Located at Both Ends of the Cluster

1999 ◽  
Vol 181 (2) ◽  
pp. 642-647 ◽  
Author(s):  
Felipe Lombó ◽  
Alfredo F. Braña ◽  
Carmen Méndez ◽  
José A. Salas
Keyword(s):  
Gene Cluster ◽  
Dna Sequences ◽  
Streptomyces Coelicolor ◽  
Regulatory Gene ◽  
Regulatory Protein ◽  
Fold Increase ◽  
Open Reading Frames ◽  
Antibiotic Biosynthesis ◽  
Positive Regulators ◽  
Actinorhodin Production

ABSTRACT Sequencing of a 4.3-kb DNA region from the chromosome ofStreptomyces argillaceus, a mithramycin producer, revealed the presence of two open reading frames (ORFs). The first one (orfA) codes for a protein that resembles several transport proteins. The second one (mtmR) codes for a protein similar to positive regulators involved in antibiotic biosynthesis (DnrI, SnoA, ActII-orf4, CcaR, and RedD) belonging to the Streptomycesantibiotic regulatory protein (SARP) family. Both ORFs are separated by a 1.9-kb, apparently noncoding region. Replacement of themtmR region by an antibiotic resistance cassette completely abolished mithramycin biosynthesis. Expression of mtmR in a high-copy-number vector in S. argillaceus caused a 16-fold increase in mithramycin production. The mtmR gene restored actinorhodin production in Streptomyces coelicolor JF1 mutant, in which the actinorhodin-specific activator ActII-orf4 is inactive, and also stimulated actinorhodin production byStreptomyces lividans TK21. A 241-bp region located 1.9 kb upstream of mtmR was found to be repeated approximately 50 kb downstream of mtmR at the other end of the mithramycin gene cluster. A model to explain a possible route for the acquisition of the mithramycin gene cluster by S. argillaceus is proposed.

Strongly Coupled Redox-Linked Conformational Switching at the Active Site of the Non-Heme Iron-Dependent Dioxygenase, TauD

2019 ◽  
Author(s):  
Christopher John ◽  
Greg M. Swain ◽  
Robert P. Hausinger ◽  
Denis A. Proshlyakov
Keyword(s):  
Active Site ◽  
Structural Model ◽  
Heme Iron ◽  
Chemical Transformations ◽  
Experimental Conditions ◽  
Strongly Coupled ◽  
Non Heme Iron ◽  
Reversible Redox ◽  
Wide Range ◽  
Complex Structural

2-Oxoglutarate (2OG)-dependent dioxygenases catalyze C-H activation while performing a wide range of chemical transformations. In contrast to their heme analogues, non-heme iron centers afford greater structural flexibility with important implications for their diverse catalytic mechanisms. We characterize an <i>in situ</i> structural model of the putative transient ferric intermediate of 2OG:taurine dioxygenase (TauD) by using a combination of spectroelectrochemical and semi-empirical computational methods, demonstrating that the Fe (III/II) transition involves a substantial, fully reversible, redox-linked conformational change at the active site. This rearrangement alters the apparent redox potential of the active site between -127 mV for reduction of the ferric state and 171 mV for oxidation of the ferrous state of the 2OG-Fe-TauD complex. Structural perturbations exhibit limited sensitivity to mediator concentrations and potential pulse duration. Similar changes were observed in the Fe-TauD and taurine-2OG-Fe-TauD complexes, thus attributing the reorganization to the protein moiety rather than the cosubstrates. Redox difference infrared spectra indicate a reorganization of the protein backbone in addition to the involvement of carboxylate and histidine ligands. Quantitative modeling of the transient redox response using two alternative reaction schemes across a variety of experimental conditions strongly supports the proposal for intrinsic protein reorganization as the origin of the experimental observations.

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