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Gating NO Release from Nitric Oxide Synthase.We have investigated the kinetics of NO escape from Geobacillus stearothermophilus nitric oxide synthase (gsNOS). Previous work indicated that NO release was gated at position 223 in mammalian enzymes; our kinetics experiments include mutants at that position along with measurements on the wild type enzyme. Employing stopped-flow UV-vis methods, reactions were triggered by mixing a reduced enzyme/N-hydroxy-l-arginine complex with an aerated buffer solution. NO release kinetics were obtained for ...201122148177
a novel cold-adapted lipase from sorangium cellulosum strain so0157-2: gene cloning, expression, and enzymatic characterization.genome sequencing of cellulolytic myxobacterium sorangium cellulosum reveals many open-reading frames (orfs) encoding various degradation enzymes with low sequence similarity to those reported, but none of them has been characterized. in this paper, a predicted lipase gene (lipa) was cloned from s. cellulosum strain so0157-2 and characterized. lipa is 981-bp in size, encoding a polypeptide of 326 amino acids that contains the pentapeptide (ghsmg) and catalytic triad residues (ser114, asp250 and ...201122072918
influence of heme-thiolate in shaping the catalytic properties of a bacterial nitric-oxide synthase.nitric-oxide synthases (nos) are heme-thiolate enzymes that generate nitric oxide (no) from l-arginine. mammalian and bacterial noss contain a conserved tryptophan (trp) that hydrogen bonds with the heme-thiolate ligand. we mutated trp(66) to his and phe (w66h, w66f) in b. subtilis nos to investigate how heme-thiolate electronic properties control enzyme catalysis. the mutations had opposite effects on heme midpoint potential (-302, -361, and -427 mv for w66h, wild-type (wt), and w66f, respectiv ...201121921039
phase i dose escalation study of kos-1584, a novel epothilone, in patients with advanced solid tumors.first-in-man study of kos-1584, a second generation epothilone.201121874318
phase i dose escalation study of kos-1584, a novel epothilone, in patients with advanced solid tumors.first-in-man study of kos-1584, a second generation epothilone.201121874318
emergence and modular evolution of a novel motility machinery in bacteria.bacteria glide across solid surfaces by mechanisms that have remained largely mysterious despite decades of research. in the deltaproteobacterium myxococcus xanthus, this locomotion allows the formation stress-resistant fruiting bodies where sporulation takes place. however, despite the large number of genes identified as important for gliding, no specific machinery has been identified so far, hampering in-depth investigations. based on the premise that components of the gliding machinery must h ...201121931562
c6-c8 bridged epothilones: consequences of installing a conformational lock at the edge of the macrocycle.a series of conformationally restrained epothilone analogues with a short bridge between the methyl groups at c6 and c8 was designed to mimic the binding pose assigned to our recently reported epoa-microtubule binding model. a versatile synthetic route to these bridged epothilone analogues has been successfully devised and implemented. biological evaluation of the compounds against a2780 human ovarian cancer and pc3 prostate cancer cell lines suggested that the introduction of a bridge between c ...201122127984
complete genome sequence of haliangium ochraceum type strain (smp-2).haliangium ochraceum fudou et al. 2002 is the type species of the genus haliangium in the myxococcal family 'haliangiaceae'. members of the genus haliangium are the first halophilic myxobacterial taxa described. the cells of the species follow a multicellular lifestyle in highly organized biofilms, called swarms, they decompose bacterial and yeast cells as most myxobacteria do. the fruiting bodies contain particularly small coccoid myxospores. h. ochraceum encodes the first actin homologue ident ...201021304682
prokaryote-derived protein inhibitors of peptidases: a sketchy occurrence and mostly unknown function.in metazoan organisms protein inhibitors of peptidases are important factors essential for regulation of proteolytic activity. in vertebrates genes encoding peptidase inhibitors constitute up to 1% of genes reflecting a need for tight and specific control of proteolysis especially in extracellular body fluids. in stark contrast unicellular organisms, both prokaryotic and eukaryotic consistently contain only few, if any, genes coding for putative peptidase inhibitors. this may seem perplexing in ...201020558234
identifying a resistance determinant for the antimitotic natural products disorazole c1 and a1.disorazoles are macrocyclic polyketides first isolated from the fermentation broth of the myxobacterium sorangium cellulosum. both the major fermentation product disorazole a(1) and its much rarer companion disorazole c(1) exhibit potent cytotoxic activity against many human tumor cells. furthermore, the disorazoles appear to bind tubulin uniquely among known antimitotic agents, promoting apoptosis or premature senescence. it is uncertain what conveys tumor cell sensitivity to these complex natu ...201020008956
biosynthesis of rhizocticins, antifungal phosphonate oligopeptides produced by bacillus subtilis atcc6633.rhizocticins are phosphonate oligopeptide antibiotics containing the c-terminal nonproteinogenic amino acid (z)-l-2-amino-5-phosphono-3-pentenoic acid (appa). here we report the identification and characterization of the rhizocticin biosynthetic gene cluster (rhi) in bacillus subtilis atcc6633. rhizocticin b was heterologously produced in the nonproducer strain bacillus subtilis 168. a biosynthetic pathway is proposed on the basis of bioinformatics analysis of the rhi genes. one of the steps dur ...201020142038
sequence-based analysis of secondary-metabolite biosynthesis in marine actinobacteria.a diverse collection of 60 marine-sediment-derived actinobacteria representing 52 operational taxonomic units was screened by pcr for genes associated with secondary-metabolite biosynthesis. three primer sets were employed to specifically target adenylation domains associated with nonribosomal peptide synthetases (nrpss) and ketosynthase (ks) domains associated with type i modular, iterative, hybrid, and enediyne polyketide synthases (pkss). in total, two-thirds of the strains yielded a sequence ...201020154113
in silico characterization of pectate lyase protein sequences from different source organisms.a total of 121 protein sequences of pectate lyases were subjected to homology search, multiple sequence alignment, phylogenetic tree construction, and motif analysis. the phylogenetic tree constructed revealed different clusters based on different source organisms representing bacterial, fungal, plant, and nematode pectate lyases. the multiple accessions of bacterial, fungal, nematode, and plant pectate lyase protein sequences were placed closely revealing a sequence level similarity. the multip ...201021048874
presence of a classical rrm-fold palm domain in thg1-type 3'- 5'nucleic acid polymerases and the origin of the ggdef and crispr polymerase domains.almost all known nucleic acid polymerases catalyze 5'-3' polymerization by mediating the attack on an incoming nucleotide 5' triphosphate by the 3'oh from the growing polynucleotide chain in a template dependent or independent manner. the only known exception to this rule is the thg1 rna polymerase that catalyzes 3'-5' polymerization in vitro and also in vivo as a part of the maturation process of histidinyl trna. while the initial reaction catalyzed by thg1 has been compared to adenylation cata ...201020591188
distinguishing microbial genome fragments based on their composition: evolutionary and comparative genomic perspectives.it is well known that patterns of nucleotide composition vary within and among genomes, although the reasons why these variations exist are not completely understood. between-genome compositional variation has been exploited to assign environmental shotgun sequences to their most likely originating genomes, whereas within-genome variation has been used to identify recently acquired genetic material such as pathogenicity islands. recent sequence assignment techniques have achieved high levels of ...201020333228
tertiary structure and characterization of a glycoside hydrolase family 44 endoglucanase from clostridium acetobutylicum.a gene encoding a glycoside hydrolase family 44 (gh44) protein from clostridium acetobutylicum atcc 824 was synthesized and transformed into escherichia coli. the previously uncharacterized protein was expressed with a c-terminal his tag and purified by nickel-nitrilotriacetic acid affinity chromatography. crystallization and x-ray diffraction to a 2.2-a resolution revealed a triose phosphate isomerase (tim) barrel-like structure with additional greek key and beta-sandwich folds, similar to othe ...201019915043
tertiary structure and characterization of a glycoside hydrolase family 44 endoglucanase from clostridium acetobutylicum.a gene encoding a glycoside hydrolase family 44 (gh44) protein from clostridium acetobutylicum atcc 824 was synthesized and transformed into escherichia coli. the previously uncharacterized protein was expressed with a c-terminal his tag and purified by nickel-nitrilotriacetic acid affinity chromatography. crystallization and x-ray diffraction to a 2.2-a resolution revealed a triose phosphate isomerase (tim) barrel-like structure with additional greek key and beta-sandwich folds, similar to othe ...201019915043
cdnl, a member of the large card-like family of bacterial proteins, is vital for myxococcus xanthus and differs functionally from the global transcriptional regulator card.card, a global transcriptional regulator in myxococcus xanthus, interacts with carg via cardnter, its n-terminal domain, and with dna via a eukaryotic hmga-type c-terminal domain. genomic analysis reveals a large number of standalone proteins resembling cardnter. these constitute, together with the rna polymerase (rnap) interacting domain, rid, of transcription-repair coupling factors, the card_trcf protein family. we show that one such cardnter-like protein, m. xanthus cdnl, cannot functionally ...201020371514
regio- and stereospecificity of filipin hydroxylation sites revealed by crystal structures of cytochrome p450 105p1 and 105d6 from streptomyces avermitilis.the polyene macrolide antibiotic filipin is widely used as a probe for cholesterol and a diagnostic tool for type c niemann-pick disease. two position-specific p450 enzymes are involved in the post-polyketide modification of filipin during its biosynthesis, thereby providing molecular diversity to the "filipin complex." cyp105p1 and cyp105d6 from streptomyces avermitilis, despite their high sequence similarities, catalyze filipin hydroxylation at different positions, c26 and c1', respectively. h ...201020375018
identification of diverse antimicrobial resistance determinants carried on bacterial, plasmid, or viral metagenomes from an activated sludge microbial assemblage.using both sequence- and function-based metagenomic approaches, multiple antibiotic resistance determinants were identified within metagenomic libraries constructed from dna extracted from bacterial chromosomes, plasmids, or viruses within an activated sludge microbial assemblage. metagenomic clones and a plasmid that in escherichia coli expressed resistance to chloramphenicol, ampicillin, or kanamycin were isolated, with many cloned dna sequences lacking any significant homology to known antibi ...201020382816
phenolic acids act as signaling molecules in plant-microbe symbioses.phenolic acids are the main polyphenols made by plants. these compounds have diverse functions and are immensely important in plant-microbe interactions/symbiosis. phenolic compounds act as signaling molecules in the initiation of legumerhizobia symbioses, establishment of arbuscular mycorrhizal symbioses and can act as agents in plant defense. flavonoids are a diverse class of polyphenolic compounds that have received considerable attention as signaling molecules involved in plant-microbe inter ...201020400851
oxazolomycin biosynthesis in streptomyces albus ja3453 featuring an "acyltransferase-less" type i polyketide synthase that incorporates two distinct extender units.the oxazolomycins (ozms) are a growing family of antibiotics produced by several streptomyces species that show diverse and important antibacterial, antitumor, and anti-human immunodeficiency virus activity. oxazolomycin a is a peptide-polyketide hybrid compound containing a unique spiro-linked beta-lactone/gamma-lactam, a 5-substituted oxazole ring. the oxazolomycin biosynthetic gene cluster (ozm) was identified from streptomyces albus ja3453 and localized to 79.5-kb dna, consisting of 20 open ...201020406823
a multi-protein complex from myxococcus xanthus required for bacterial gliding motility.myxococcus xanthus moves by gliding motility powered by type iv pili (s-motility) and a second motility system, a-motility, whose mechanism remains elusive despite the identification of approximately 40 a-motility genes. in this study, we used biochemistry and cell biology analyses to identify multi-protein complexes associated with a-motility. previously, we showed that the n-terminal domain of frzcd, the receptor for the frizzy chemosensory pathway, interacts with two a-motility proteins, aglz ...201020487265
using chemobiosynthesis and synthetic mini-polyketide synthases to produce pharmaceutical intermediates in escherichia coli.recombinant microbial whole-cell biocatalysis is a valuable approach for producing enantiomerically pure intermediates for the synthesis of complex molecules. here, we describe a method to produce polyketide intermediates possessing multiple stereogenic centers by combining chemobiosynthesis and engineered mini-polyketide synthases (pkss). chemobiosynthesis allows the introduction of unnatural moieties, while a library of synthetic bimodular pkss expressed from codon-optimized genes permits the ...201020543042
the phosphatomes of the multicellular myxobacteria myxococcus xanthus and sorangium cellulosum in comparison with other prokaryotic genomes.analysis of the complete genomes from the multicellular myxobacteria myxococcus xanthus and sorangium cellulosum identified the highest number of eukaryotic-like protein kinases (elks) compared to all other genomes analyzed. high numbers of protein phosphatases (pps) could therefore be anticipated, as reversible protein phosphorylation is a major regulation mechanism of fundamental biological processes.201020567509
the bridge-region of the ku superfamily is an atypical zinc ribbon domain.members of the ku superfamily are dna-end-binding proteins involved in non-homologous end-joining (nhej) dna repair. the published crystal structure of human ku-dna complex reveals a heterodimer that forms a ring around dsdna by means of the ku core modules. these modules contain a highly conserved seven-stranded β-barrel, which in turn contains an insertion, termed the bridge-region, between its second and third β-strands. the bridge-region adopts an unusual β-strand-rich structure critical for ...201020580930
discovery and total synthesis of a new estrogen receptor heterodimerizing actinopolymorphol a from actinopolymorpha rutilus.estrogen receptor eralpha and erbeta heterodimerization has been implicated in cancer chemoprevention. the discovery, structural elucidation, and total synthesis of a new natural product, actinopolymorphol a (1), from actinopolymorpha rutilus (yim45725) that preferentially induces eralpha/beta heterodimerization is reported. total synthesis of 1 has allowed us to determine its absolute stereochemistry and that of a previously known deacetylated congener, and 1 represents the first member of a ne ...201020593804
structural insights into the function of aminoglycoside-resistance a1408 16s rrna methyltransferases from antibiotic-producing and human pathogenic bacteria.x-ray crystal structures were determined of the broad-spectrum aminoglycoside-resistance a1408 16s rrna methyltransferases kamb and npma, from the aminoglycoside-producer streptoalloteichus tenebrarius and human pathogenic escherichia coli, respectively. consistent with their common function, both are class i methyltransferases with additional highly conserved structural motifs that embellish the core sam-binding fold. in overall structure, the a1408 rrna methyltransferase were found to be most ...201020639535
from complete genome sequence to 'complete' understanding?the rapidly accumulating genome sequence data allow researchers to address fundamental biological questions that were not even asked just a few years ago. a major problem in genomics is the widening gap between the rapid progress in genome sequencing and the comparatively slow progress in the functional characterization of sequenced genomes. here we discuss two key questions of genome biology: whether we need more genomes, and how deep is our understanding of biology based on genomic analysis. w ...201020647113
damage of streptococcus mutans biofilms by carolacton, a secondary metabolite from the myxobacterium sorangium cellulosum.streptococcus mutans is a major pathogen in human dental caries. one of its important virulence properties is the ability to form biofilms (dental plaque) on tooth surfaces. eradication of such biofilms is extremely difficult. we therefore screened a library of secondary metabolites from myxobacteria for their ability to damage biofilms of s. mutans.201020659313
identification of the biosynthetic gene cluster for the pacidamycin group of peptidyl nucleoside antibiotics.pacidamycins are a family of uridyl tetra/pentapeptide antibiotics that act on the translocase mray to block bacterial cell wall assembly. to elucidate the biosynthetic logic of pacidamcyins, a putative gene cluster was identified by 454 shotgun genome sequencing of the producer streptomyces coeruleorubidus nrrl 18370. the 31-kb gene cluster encodes 22 proteins (paca-v), including highly dissociated nonribosomal peptide synthetase (nrps) modules and a variety of tailoring enzymes. gene deletions ...201020826445
development of an environmental functional gene microarray for soil microbial communities.functional attributes of microbial communities are difficult to study, and most current techniques rely on dna- and rrna-based profiling of taxa and genes, including microarrays containing sequences of known microorganisms. to quantify gene expression in environmental samples in a culture-independent manner, we constructed an environmental functional gene microarray (e-fga) consisting of 13,056 mrna-enriched anonymous microbial clones from diverse microbial communities to profile microbial gene ...201020851978
engineered polyketide biosynthesis and biocatalysis in escherichia coli.polyketides are important bioactive natural products biosynthesized by bacteria, fungi, and plants. the enzymes that synthesize polyketides are collectively referred to as polyketide synthases (pkss). because many of the natural hosts that produce polyketides are difficult to culture or manipulate, establishing a universal heterologous host that is genetically tractable has become an important goal toward the engineered biosynthesis of polyketides and analogues. here, we summarize the recent pro ...201020853106
structural and biochemical characterization of mycobacterium tuberculosis cyp142: evidence for multiple cholesterol 27-hydroxylase activities in a human pathogen.the mycobacterium tuberculosis cytochrome p450 enzyme cyp142 is encoded in a large gene cluster involved in metabolism of host cholesterol. cyp142 was expressed and purified as a soluble, low spin p450 hemoprotein. cyp142 binds tightly to cholesterol and its oxidized derivative cholest-4-en-3-one, with extensive shift of the heme iron to the high spin state. high affinity for azole antibiotics was demonstrated, highlighting their therapeutic potential. cyp142 catalyzes either 27-hydroxylation of ...201020889498
marine myxobacteria as a source of antibiotics--comparison of physiology, polyketide-type genes and antibiotic production of three new isolates of enhygromyxa salina.three myxobacterial strains, designated swb004, swb005 and swb006, were obtained from beach sand samples from the pacific ocean and the north sea. the strains were cultivated in salt water containing media and subjected to studies to determine their taxonomic status, the presence of genes for the biosynthesis of polyketides and antibiotic production. 16s rdna sequence analysis revealed the type strain enhygromyxa salina shk-1(t) as their closest homolog, displaying between 98% (swb005) and 99% ( ...201020948900
effects of site-directed mutagenesis of mgla on motility and swarming of myxococcus xanthus.the mgla gene from the bacterium myxococcus xanthus encodes a 22kda protein related to the ras superfamily of monomeric gtpases. mgla is required for the normal function of a-motility (adventurous), s-motility (social), fruiting body morphogenesis, and sporulation. mgla and its homologs differ from all eukaryotic and other prokaryotic gtpases because they have a threonine (thr78) in place of the highly conserved aspartate residue of the consensus pm3 (phosphate-magnesium binding) region. to iden ...201021083931
evolution of bacterial phosphoglycerate mutases: non-homologous isofunctional enzymes undergoing gene losses, gains and lateral transfers.the glycolytic phosphoglycerate mutases exist as non-homologous isofunctional enzymes (nise) having independent evolutionary origins and no similarity in primary sequence, 3d structure, or catalytic mechanism. cofactor-dependent pgm (dpgm) requires 2,3-bisphosphoglycerate for activity; cofactor-independent pgm (ipgm) does not. the pgm profile of any given bacterium is unpredictable and some organisms such as escherichia coli encode both forms.201021187861
tir domain-containing adaptor sarm is a late addition to the ongoing microbe-host dialog.toll/interleukin-1 receptor (tir) domain-containing proteins play important roles in defense against pathogens in both animals and plants, connecting the immunity signaling pathways via a chain of specific protein-protein interactions. among them is sarm, the only tir domain-containing adaptor that can negatively regulate tlr signaling. by extensive phylogenetic analysis, we show here that sarm is closely related to bacterial proteins with tir domains, suggesting that this family has a different ...201021110998
tir domain-containing adaptor sarm is a late addition to the ongoing microbe-host dialog.toll/interleukin-1 receptor (tir) domain-containing proteins play important roles in defense against pathogens in both animals and plants, connecting the immunity signaling pathways via a chain of specific protein-protein interactions. among them is sarm, the only tir domain-containing adaptor that can negatively regulate tlr signaling. by extensive phylogenetic analysis, we show here that sarm is closely related to bacterial proteins with tir domains, suggesting that this family has a different ...201021110998
networks of gene sharing among 329 proteobacterial genomes reveal differences in lateral gene transfer frequency at different phylogenetic depths.lateral gene transfer (lgt) is an important mechanism of natural variation among prokaryotes. over the full course of evolution, most or all of the genes resident in a given prokaryotic genome have been affected by lgt, yet the frequency of lgt can vary greatly across genes and across prokaryotic groups. the proteobacteria are among the most diverse of prokaryotic taxa. the prevalence of lgt in their genome evolution calls for the application of network-based methods instead of tree-based method ...201021059789
networks of gene sharing among 329 proteobacterial genomes reveal differences in lateral gene transfer frequency at different phylogenetic depths.lateral gene transfer (lgt) is an important mechanism of natural variation among prokaryotes. over the full course of evolution, most or all of the genes resident in a given prokaryotic genome have been affected by lgt, yet the frequency of lgt can vary greatly across genes and across prokaryotic groups. the proteobacteria are among the most diverse of prokaryotic taxa. the prevalence of lgt in their genome evolution calls for the application of network-based methods instead of tree-based method ...201021059789
molecular evolution and the role of oxidative stress in the expansion and functional diversification of cytosolic glutathione transferases.cytosolic glutathione transferases (cgst) are a large group of ubiquitous enzymes involved in detoxification and are well known for their undesired side effects during chemotherapy. in this work we have performed thorough phylogenetic analyses to understand the various aspects of the evolution and functional diversification of cgsts. furthermore, we assessed plausible correlations between gene duplication and substrate specificity of gene paralogs in humans and selected species, notably in mamma ...201020843339
myxobacteria, polarity, and multicellular morphogenesis.myxobacteria are renowned for the ability to sporulate within fruiting bodies whose shapes are species-specific. the capacity to build those multicellular structures arises from the ability of m. xanthus to organize high cell-density swarms, in which the cells tend to be aligned with each other while constantly in motion. the intrinsic polarity of rod-shaped cells lays the foundation, and each cell uses two polar engines for gliding on surfaces. it sprouts retractile type iv pili from the leadin ...201020610548
chemotherapy resistance in metastatic breast cancer: the evolving role of ixabepilone.resistance to chemotherapy is a major obstacle to the effective treatment of many tumor types. although many anticancer therapies can alter tumor growth, in most cases the effect is not long lasting. consequently, there is a significant need for new agents with low susceptibility to common drug resistance mechanisms in order to improve response rates and potentially extend survival. approximately 30% of the women diagnosed with early-stage disease in turn progress to metastatic breast cancer, fo ...201021050423
prediction and analysis of the modular structure of cytochrome p450 monooxygenases.cytochrome p450 monooxygenases (cyps) form a vast and diverse family of highly variable sequences. they catalyze a wide variety of oxidative reactions and are therefore of great relevance in drug development and biotechnological applications. despite their differences in sequence and substrate specificity, the structures of cyps are highly similar. although being in research focus for years, factors mediating selectivity and activity remain vague.201020950472
conformational plasticity and structure/function relationships in cytochromes p450.the cytochrome p450s are a superfamily of enzymes that are found in all kingdoms of living organisms, and typically catalyze the oxidative addition of atomic oxygen to an unactivated c-c or c-h bond. over 8000 nonredundant sequences of putative and confirmed p450 enzymes have been identified, but three-dimensional structures have been determined for only a small fraction of these. while all p450 enzymes for which structures have been determined share a common global fold, the flexibility and mod ...201020446763
endometrial carcinoma: a review of chemotherapy, drug resistance, and the search for new agents.adenocarcinoma of the endometrium represents the most common gynecologic malignancy in developed countries. although early-stage cancers are effectively treated surgically, commonly without adjuvant therapy, the treatment of high-risk and advanced disease is more complex. chemotherapy has evolved into an important modality in high-risk early-stage and advanced-stage disease, and in recurrent endometrial cancer. taxane-based therapy consistently demonstrates the highest response rates in the firs ...201020930101
microtubule-binding agents: a dynamic field of cancer therapeutics.microtubules are dynamic filamentous cytoskeletal proteins composed of tubulin and are an important therapeutic target in tumour cells. agents that bind to microtubules have been part of the pharmacopoeia of anticancer therapy for decades and until the advent of targeted therapy, microtubules were the only alternative to dna as a therapeutic target in cancer. the screening of a range of botanical species and marine organisms has yielded promising new antitubulin agents with novel properties. in ...201020885410
optimizing ixabepilone treatment schedules in patients with advanced or metastatic breast cancer.the epothilone b analog, ixabepilone, demonstrates low susceptibility to drug resistance mechanisms and has demonstrated clinically meaningful efficacy in patients refractory to other chemotherapeutic options. ixabepilone is approved by the fda for treatment of patients with metastatic breast cancer (mbc) progressing after taxanes and anthracyclines, either in combination with capecitabine or as monotherapy if the patient has already progressed on capecitabine. ixabepilone is generally well tole ...201020886213
detecting and understanding the roles of nitric oxide in biology.we are pursuing a dual strategy for investigating the chemistry of nitric oxide as a biological signaling agent. in one approach, metal-based fluorescent sensors for the detection of no in living cells are evaluated, and a sensor based on a copper fluorescein complex has proved to be a valuable lead compound. sensors of this class permit identification of no from both inducible and constitutive forms of nitric oxide synthase and facilitate investigation of different no functions in response to e ...201020666391
read length and repeat resolution: exploring prokaryote genomes using next-generation sequencing technologies.there are a growing number of next-generation sequencing technologies. at present, the most cost-effective options also produce the shortest reads. however, even for prokaryotes, there is uncertainty concerning the utility of these technologies for the de novo assembly of complete genomes. this reflects an expectation that short reads will be unable to resolve small, but presumably abundant, repeats.201020634954
identification of the viridicatumtoxin and griseofulvin gene clusters from penicillium aethiopicum.penicillium aethiopicum produces two structurally interesting and biologically active polyketides: the tetracycline-like viridicatumtoxin 1 and the classic antifungal agent griseofulvin 2. here, we report the concurrent discovery of the two corresponding biosynthetic gene clusters (vrt and gsf) by 454 shotgun sequencing. gene deletions confirmed that two nonreducing pkss (nrpkss), vrta and gsfa, are required for the biosynthesis of 1 and 2, respectively. both pkss share similar domain architectu ...201020534346
ixabepilone as monotherapy or in combination for the treatment of advanced breast cancer.advanced breast cancer represents a therapeutic challenge for oncologists. chemotherapy with anthracyclines and taxanes has improved survival in this setting, but resistant and refractory disease is common. ixabepilone, an epothilone, is a recently approved chemotherapeutic agent, both as a single agent and in combination with capecitabine, with efficacy in patients with resistant advanced disease. in this review, the distinctive properties of this drug are discussed, as well as the clinical evi ...201024367163
the mixed lineage nature of nitrogen transport and assimilation in marine eukaryotic phytoplankton: a case study of micromonas.the prasinophyte order mamiellales contains several widespread marine picophytoplankton (≤ 2 μm diameter) taxa, including micromonas and ostreococcus. complete genome sequences are available for two micromonas isolates, ccmp1545 and rcc299. we performed in silico analyses of nitrogen transporters and related assimilation genes in ccmp1545 and rcc299 and compared these with other green lineage organisms as well as chromalveolata, fungi, bacteria, and archaea. phylogenetic reconstructions of ammon ...201020457585
a phase i/ii trial and pharmacokinetic study of ixabepilone in adult patients with recurrent high-grade gliomas.ixabepilone is an epothilone, a novel class of non-taxane microtubule stabilizing agents. a phase i/ii and pharmacokinetic trial of ixabepilone was conducted in patients with recurrent high-grade gliomas. adult patients received ixabepilone as a 1-h infusion daily for 5 days every 3 weeks. a modified continual reassessment method was used to escalate doses, beginning at 5.0 mg/m(2), in patients stratified by use or non-use of enzyme inducing antiepileptic drugs (eiaed). in the phase i study, the ...201020449631
recognition of (2s)-aminomalonyl-acyl carrier protein (acp) and (2r)-hydroxymalonyl-acp by acyltransferases in zwittermicin a biosynthesis.polyketide synthases elongate a polyketide backbone by condensing carboxylic acid precursors that are thioesterified to either coenzyme a or an acyl carrier protein (acp). two of the three known acp-linked extender units, (2s)-aminomalonyl-acp and (2r)-hydroxymalonyl-acp, are found in the biosynthesis of the agriculturally important antibiotic zwittermicin a. we previously reconstituted the formation of (2s)-aminomalonyl-acp and (2r)-hydroxymalonyl-acp from the primary metabolites l-serine and 1 ...201020353188
phase i trial of ixabepilone plus pegylated liposomal doxorubicin in patients with adenocarcinoma of breast or ovary.ixabepilone is a semisynthetic epothilone b analogue that is active in taxane-resistant cell lines and has shown activity in patients with refractory breast and ovarian cancer. we carried out a phase i trial of ixabepilone plus pegylated liposomal doxorubicin (pld) in patients with advanced taxane-pretreated ovarian and breast cancer.201020357034
mitotic slippage in non-cancer cells induced by a microtubule disruptor, disorazole c1.disorazoles are polyene macrodiolides isolated from a myxobacterium fermentation broth. disorazole c1 was newly synthesized and found to depolymerize microtubules and cause mitotic arrest. here we examined the cellular responses to disorazole c1 in both non-cancer and cancer cells and compared our results to vinblastine and taxol.201020181182
a phase ii trial of the epothilone b analog ixabepilone (bms-247550) in patients with metastatic melanoma.ixabepilone (bms-247550), an epothilone b analog, is a microtubule stabilizing agent which has shown activity in several different tumor types and preclinical models in melanoma. in an open label, one-arm, multi-center phase ii trial the efficacy and toxicity of this epothilone was investigated in two different cohorts: chemotherapy-naïve (previously untreated) and previously treated patients with metastatic melanoma.201020098694
total synthesis and evaluation of c26-hydroxyepothilone d derivatives for photoaffinity labeling of beta-tubulin.three photoaffinity labeled derivatives of epothilone d were prepared by total synthesis, using efficient novel asymmetric synthesis methods for the preparation of two important synthetic building blocks. the key step for the asymmetric synthesis of (s,e)-3-(tert-butyldimethylsilyloxy)-4-methyl-5-(2-methylthiazol-4-yl)pent-4-enal involved a ketone reduction with (r)-me-cbs-oxazaborolidine. for the synthesis of (5s)-5,7-di[(tert-butyldimethylsilyl)oxy]-4,4-dimethylheptan-3-one an asymmetric noyor ...201019954175
type i polyketide synthases that require discrete acyltransferases.the diverse structures of polyketide natural products are reflected by the equally diverse polyketide biosynthetic enzymes, namely polyketide synthases (pkss). three major classes of pkss are known-noniterative type i pkss, iterative type ii pkss and acyl carrier protein-independent type iii pkss, each of which consists of additional variants. one such variant is the noniterative type i pks in which each pks module lacks the cognate acyltransferase (at) domain. the essential at activity is inste ...200919362640
microbial-based therapy of cancer: current progress and future prospects.the use of bacteria in the regression of certain forms of cancer has been recognized for more than a century. much effort, therefore, has been spent over the years in developing wild-type or modified bacterial strains to treat cancer. however, their use at the dose required for therapeutic efficacy has always been associated with toxicity problems and other deleterious effects. recently, the old idea of using bacteria in the treatment of cancer has attracted considerable interest and new genetic ...200921326924
microbial-based therapy of cancer: current progress and future prospects.the use of bacteria in the regression of certain forms of cancer has been recognized for more than a century. much effort, therefore, has been spent over the years in developing wild-type or modified bacterial strains to treat cancer. however, their use at the dose required for therapeutic efficacy has always been associated with toxicity problems and other deleterious effects. recently, the old idea of using bacteria in the treatment of cancer has attracted considerable interest and new genetic ...200921326924
structures of rna polymerase-antibiotic complexes.inhibition of bacterial rna polymerase (rnap) is an established strategy for antituberculosis therapy and broad-spectrum antibacterial therapy. crystal structures of rnap-inhibitor complexes are available for four classes of antibiotics: rifamycins, sorangicin, streptolydigin, and myxopyronin. the structures define three different targets, and three different mechanisms, for inhibition of bacterial rnap: (1) rifamycins and sorangicin bind near the rnap active center and block extension of rna pr ...200919926275
recombinant organisms for production of industrial products.a revolution in industrial microbiology was sparked by the discoveries of ther double-stranded structure of dna and the development of recombinant dna technology. traditional industrial microbiology was merged with molecular biology to yield improved recombinant processes for the industrial production of primary and secondary metabolites, protein biopharmaceuticals and industrial enzymes. novel genetic techniques such as metabolic engineering, combinatorial biosynthesis and molecular breeding te ...200921326937
recombinant organisms for production of industrial products.a revolution in industrial microbiology was sparked by the discoveries of ther double-stranded structure of dna and the development of recombinant dna technology. traditional industrial microbiology was merged with molecular biology to yield improved recombinant processes for the industrial production of primary and secondary metabolites, protein biopharmaceuticals and industrial enzymes. novel genetic techniques such as metabolic engineering, combinatorial biosynthesis and molecular breeding te ...200921326937
a subset of the diverse cog0523 family of putative metal chaperones is linked to zinc homeostasis in all kingdoms of life.cog0523 proteins are, like the nickel chaperones of the ureg family, part of the g3e family of gtpases linking them to metallocenter biosynthesis. even though the first cog0523-encoding gene, cobw, was identified almost 20 years ago, little is known concerning the function of other members belonging to this ubiquitous family.200919822009
total synthesis of (+)-sorangicin a.the final synthetic challenges associated with (+)-sorangicin a have been overcome, thus leading to the first total synthesis of this complex macrolide antibiotic. highlights of the highly convergent synthesis include two julia-kociénski olefinations to unite three advanced fragments with high e-stereoselectivity. critical to the final-stage success was the use of a carefully defined stille coupling and a mukaiyama macrolactonization as well as lewis and protic acid-promoted deprotections carefu ...200919663510
the 1.5-a structure of xpla-heme, an unusual cytochrome p450 heme domain that catalyzes reductive biotransformation of royal demolition explosive.xpla is a cytochrome p450 of unique structural organization, consisting of a heme-domain that is c-terminally fused to its native flavodoxin redox partner. xpla, along with flavodoxin reductase xplb, has been shown to catalyze the breakdown of the nitramine explosive and pollutant hexahydro-1,3,5-trinitro-1,3,5-triazine (royal demolition explosive) by reductive denitration. the structure of the heme domain of xpla (xpla-heme) has been solved in two crystal forms: as a dimer in space group p2(1) ...200919692330
the epothilone b analogue ixabepilone in patients with advanced hepatobiliary cancers: a trial of the university of chicago phase ii consortium.hepatobiliary cancers respond poorly to cytotoxic chemotherapy. we evaluated the activity and safety of ixabepilone, an epothilone b analogue which stabilizes microtubules, in a phase ii trial in patients with advanced cancers of the gallbladder, bile duct, and liver.200919669700
the epothilone b analogue ixabepilone in patients with advanced hepatobiliary cancers: a trial of the university of chicago phase ii consortium.hepatobiliary cancers respond poorly to cytotoxic chemotherapy. we evaluated the activity and safety of ixabepilone, an epothilone b analogue which stabilizes microtubules, in a phase ii trial in patients with advanced cancers of the gallbladder, bile duct, and liver.200919669700
recent applications of acyclic (diene)iron complexes and (dienyl)iron cations in organic synthesis.complexation of (tricarbonyl)iron to an acyclic diene serves to protect the ligand against oxidation, reduction and cycloaddition reactions while the steric bulk of this adjunct serves to direct the approach reagents to unsaturated groups attached to the diene onto the face opposite to iron. furthermore, the fe(co)(3) moiety can serve to stabilize carbocation centers adjacent to the diene (i.e. pentadienyl-iron cations). recent applications of these reactivities to the synthesis of polyene, cycl ...200921709767
automated genome mining for natural products.discovery of new medicinal agents from natural sources has largely been an adventitious process based on screening of plant and microbial extracts combined with bioassay-guided identification and natural product structure elucidation. increasingly rapid and more cost-effective genome sequencing technologies coupled with advanced computational power have converged to transform this trend toward a more rational and predictive pursuit.200919531248
treatment options for breast cancer resistant to anthracycline and taxane.breast cancer is the most common noncutaneous malignancy among every major ethnic group of women in the united states. anthracyclines and taxanes are the most active and widely used chemotherapeutic agents for breast cancer, but the increased use of these agents at an early stage of disease often renders tumors resistant to these drugs by the time the disease recurs, thereby reducing the number of treatment options for metastatic disease. moreover, even when these agents can be used in the metas ...200919483170
total synthesis and evaluation of 22-(3-azidobenzoyloxy)methyl epothilone c for photoaffinity labeling of beta-tubulin.the total synthesis of 22-(3-azidobenzoyloxy)methyl epothilone c is described as a potential photoaffinity probe to elucidate the beta-tubulin binding site. a sequential suzuki-aldol-yamaguchi macrolactonization strategy was utilized employing a novel derivatized c1-c6 fragment. the c22-functionalized analog exhibited good activity in microtubule assembly assays, but cytotoxicity was significantly reduced. molecular modeling simulations indicated that excessive steric bulk in the c22 position is ...200919428248
genome degeneration affects both extracellular and intracellular bacterial endosymbionts.the obligate intracellular bacterial endosymbionts of insects are a paradigm for reductive genome evolution. a study published recently in bmc biology demonstrates that similar evolutionary forces shaping genome structure may also apply to extracellular endosymbionts.200919435469
an efficient, second-generation synthesis of the signature dioxabicyclo[3.2.1]octane core of (+)-sorangicin a and elaboration of the (z,z,e)-triene acid system.an efficient, second-generation synthesis of the signature dioxabicyclo[3.2.1]octane core of (+)-sorangicin a (1), in conjunction with an effective, stereocontrolled protocol to arrive at the requisite (z,z,e)-triene acid system has been developed. highlights of the core construction entail a three-component union, a khmds-promoted epoxide ring formation-ring opening cascade, a takai olefination, and a chemoselective sharpless dihydroxylation. assembly of the triene acid system was then achieved ...200919182995
tubulin-interactive natural products as anticancer agents.this review provides an overview of the discovery, structures, and biological activities of anticancer natural products that act by inhibiting or promoting the assembly of tubulin to microtubules. the emphasis is on providing recent information on those compounds in clinical use or in advanced clinical trials. the vinca alkaloids, the combretastatins, npi-2358, the halichondrin b analogue eribulin, dolastatin 10, noscapine, hemiasterlin, and rhizoxin are discussed as tubulin polymerization inhib ...200919125622
alterations of chemotherapeutic pharmacokinetic profiles by drug-drug interactions.drug interactions in oncology are common place and largely ignored as we tolerate high thresholds of 'toxic' drug responses in these patients. however, in the era of 'targeted' or seemingly 'less toxic' therapy, these interactions are more commonly flagged and contribute significantly towards poor 'quality of life' and medical fatalities.200919239394
asymmetric total synthesis of soraphen a: a flexible alkyne strategy. 200919554583
biosynthesis of polyketide synthase extender units.this review covers the biosynthesis of extender units that are utilized for the assembly of polyketides by polyketide synthases. the metabolic origins of each of the currently known polyketide synthase extender units are covered.200919374124
management of advanced breast cancer with the epothilone b analog, ixabepilone.despite the activity of standard chemotherapies in advanced breast cancer, disease progression remains inevitable. most patients exposed to anthracyclines and taxanes develop resistance and a significant subset shows primary resistance. the increasing use of these agents as adjuvant therapy may result in more anthracycline- and taxane-resistant patients in the metastatic setting; few treatment options are available for patients with metastatic breast cancer (mbc) resistant to multiple chemothera ...200919920932
the group iii two-component histidine kinase of filamentous fungi is involved in the fungicidal activity of the bacterial polyketide ambruticin.we have shown that the plant pathogen alternaria brassicicola exhibited very high susceptibility to ambruticin vs4 and to a lesser extent to the phenylpyrrole fungicide fludioxonil. these compounds are both derived from natural bacterial metabolites with antifungal properties and are thought to exert their toxicity by interfering with osmoregulation in filamentous fungi. disruption of the osmosensor group iii histidine kinase gene abnik1 (for a. brassicola nik1) resulted in high levels of resist ...200919011080
the group iii two-component histidine kinase of filamentous fungi is involved in the fungicidal activity of the bacterial polyketide ambruticin.we have shown that the plant pathogen alternaria brassicicola exhibited very high susceptibility to ambruticin vs4 and to a lesser extent to the phenylpyrrole fungicide fludioxonil. these compounds are both derived from natural bacterial metabolites with antifungal properties and are thought to exert their toxicity by interfering with osmoregulation in filamentous fungi. disruption of the osmosensor group iii histidine kinase gene abnik1 (for a. brassicola nik1) resulted in high levels of resist ...200919011080
microtubule binding and disruption and induction of premature senescence by disorazole c(1).disorazoles comprise a family of 29 macrocyclic polyketides isolated from the fermentation broth of the myxobacterium sorangium cellulosum. the major fermentation product, disorazole a(1), was found previously to irreversibly bind to tubulin and to have potent cytotoxic activity against tumor cells, possibly because of its highly electrophilic epoxide moiety. to test this hypothesis, we synthesized the epoxide-free disorazole c(1) and found it retained potent antiproliferative activity against t ...200919066338
microtubule binding and disruption and induction of premature senescence by disorazole c(1).disorazoles comprise a family of 29 macrocyclic polyketides isolated from the fermentation broth of the myxobacterium sorangium cellulosum. the major fermentation product, disorazole a(1), was found previously to irreversibly bind to tubulin and to have potent cytotoxic activity against tumor cells, possibly because of its highly electrophilic epoxide moiety. to test this hypothesis, we synthesized the epoxide-free disorazole c(1) and found it retained potent antiproliferative activity against t ...200919066338
inactivation of the ketoreductase gilu gene of the gilvocarcin biosynthetic gene cluster yields new analogues with partly improved biological activity.four new analogues of the gilvocarcin-type aryl-c-glycoside antitumor compounds, namely 4'-hydroxy gilvocarcin v (4'-oh-gv), 4'-hydroxy gilvocarcin m, 4'-hydroxy gilvocarcin e and 12-demethyl-defucogilvocarcin v, were produced through inactivation of the gilu gene. the 4'-oh-analogues showed improved activity against lung cancer cell lines as compared to their parent compounds without 4'-oh group (gilvocarcins v and e). the structures of the sugar-containing new mutant products indicate that the ...200919067453
novel expression hosts for complex secondary metabolite megasynthetases: production of myxochromide in the thermopilic isolate corallococcus macrosporus gt-2.abstract: although many secondary metabolites with diverse biological activities have been isolated from myxobacteria, most strains of these biotechnologically important gliding prokaryotes remain difficult to handle genetically. in this study we describe the new fast growing myxobacterial thermophilic isolate gt-2 as a heterologous host for the expression of natural product biosynthetic pathways isolated from other myxobacteria. according to the results of sequence analysis of the 16s rdna, thi ...200919126236
high-level production of amorpha-4,11-diene, a precursor of the antimalarial agent artemisinin, in escherichia coli.background: artemisinin derivatives are the key active ingredients in artemisinin combination therapies (acts), the most effective therapies available for treatment of malaria. because the raw material is extracted from plants with long growing seasons, artemisinin is often in short supply, and fermentation would be an attractive alternative production method to supplement the plant source. previous work showed that high levels of amorpha-4,11-diene, an artemisinin precursor, can be made in esch ...200919221601
formation and metabolism of methylmalonyl coenzyme a in corynebacterium glutamicum.genome sequence information suggests that b(12)-dependent mutases are present in a number of bacteria, including members of the suborder corynebacterineae like mycobacterium tuberculosis and corynebacterium glutamicum. we here functionally identify a methylmalonyl coenzyme a (coa) mutase in c. glutamicum that is retained in all of the members of the suborder corynebacterineae and is encoded by ncgl1471, ncgl1472, and ncgl1470. in addition, we observe the presence of methylmalonate in c. glutamic ...200919233926
formation and characterization of acyl carrier protein-linked polyketide synthase extender units.polyketide natural products are assembled by the condensation of an initiating precursor, or starter unit, with a series of additional precursors referred to as extender units. while there are a number of polyketide synthase starter units, there are currently only seven known polyketide synthase extender units. polyketide synthase extender units thioesterified to coenzyme a have been known for some time; however, polyketide synthase extender units thioesterified to acyl carrier proteins (acps) h ...200919362639
isolation, biology and chemistry of the disorazoles: new anti-cancer macrodiolides.covering: 1994 to 2008. the disorazoles comprise a family of 29 closely related macrocyclic polyketides isolated in 1994 from the fermentation broth of the gliding myxobacterium sorangium cellulosum. disorazoles a1, e and c1 have shown exceptional biological activites toward inhibiting the proliferation of human cancer cell lines in picomolar and nanomolar concentrations through the disruption of microtubule polymerization. this review gives a brief introduction describing the biosynthesis and t ...200919387496
measure of synonymous codon usage diversity among genes in bacteria.in many bacteria, intragenomic diversity in synonymous codon usage among genes has been reported. however, no quantitative attempt has been made to compare the diversity levels among different genomes. here, we introduce a mean dissimilarity-based index (dmean) for quantifying the level of diversity in synonymous codon usage among all genes within a genome.200919480720
insertion of horizontally transferred genes within conserved syntenic regions of yeast genomes.horizontal gene transfer has been occasionally mentioned in eukaryotic genomes, but such events appear much less numerous than in prokaryotes, where they play important functional and evolutionary roles. in yeasts, few independent cases have been described, some of which corresponding to major metabolic functions, but no systematic screening of horizontally transferred genes has been attempted so far. taking advantage of the synteny conservation among five newly sequenced and annotated genomes o ...200919654869
genome mining in sorangium cellulosum so ce56: identification and characterization of the homologous electron transfer proteins of a myxobacterial cytochrome p450.myxobacteria, especially members of the genus sorangium, are known for their biotechnological potential as producers of pharmaceutically valuable secondary metabolites. the biosynthesis of several of those myxobacterial compounds includes cytochrome p450 activity. although class i cytochrome p450 enzymes occur wide-spread in bacteria and rely on ferredoxins and ferredoxin reductases as essential electron mediators, the study of these proteins is often neglected. therefore, we decided to search i ...200919696019
myxococcus xanthus pph2 is a manganese-dependent protein phosphatase involved in energy metabolism.the multicellular behavior of the myxobacterium myxococcus xanthus requires the participation of an elevated number of signal-transduction mechanisms to coordinate the cell movements and the sequential changes in gene expression patterns that lead to the morphogenetic and differentiation events. these signal-transduction mechanisms are mainly based on two-component systems and on the reversible phosphorylation of protein targets mediated by eukaryotic-like protein kinases and phosphatases. among ...200919706604
no formation by a catalytically self-sufficient bacterial nitric oxide synthase from sorangium cellulosum.the role of nitric oxide (no) in the host response to infection and in cellular signaling is well established. enzymatic synthesis of no is catalyzed by the nitric oxide synthases (noss), which convert arg into no and citrulline using co-substrates o2 and nadph. mammalian nos contains a flavin reductase domain (fad and fmn) and a catalytic heme oxygenase domain (p450-type heme and tetrahydrobiopterin). bacterial noss, while much less studied, were previously identified as only containing the hem ...200919805284
tonb-dependent transporters and their occurrence in cyanobacteria.different iron transport systems evolved in gram-negative bacteria during evolution. most of the transport systems depend on outer membrane localized tonb-dependent transporters (tbdts), a periplasma-facing tonb protein and a plasma membrane localized machinery (exbbd). so far, iron chelators (siderophores), oligosaccharides and polypeptides have been identified as substrates of tbdts. for iron transport, three uptake systems are defined: the lactoferrin/transferrin binding proteins, the porphyr ...200919821963
p2cs: a two-component system resource for prokaryotic signal transduction research.with the escalation of high throughput prokaryotic genome sequencing, there is an ever-increasing need for databases that characterise, catalogue and present data relating to particular gene sets and genomes/metagenomes. two-component system (tcs) signal transduction pathways are the dominant mechanisms by which micro-organisms sense and respond to external as well as internal environmental changes. these systems respond to a wide range of stimuli by triggering diverse physiological adjustments, ...200919604365
metronomic dosing enhances the anti-angiogenic effect of epothilone b.background: high doses (10nm) of epothilone b, a microtubule stabilizer, will inhibit the development of human tumor-derived angiogenesis following short (14 d) drug exposure times. metronomic dosing regimes use lower drug doses and prolonged drug exposure times in an attempt to decrease toxicity compared with standard dosing schedules. we hypothesized that epothilone b would be an effective anti-angiogenic agent when administered at very low doses over an extended period of time. methods: fragm ...200920338584
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