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nitric oxide dioxygenase: an enzymic function for flavohemoglobin.nitric oxide (no*) is a toxin, and various life forms appear to have evolved strategies for its detoxification. no*-resistant mutants of escherichia coli were isolated that rapidly consumed no*. an no*-converting activity was reconstituted in extracts that required nadph, fad, and o2, was cyanide-sensitive, and produced no3-. this nitric oxide dioxygenase (nod) contained 19 of 20 n-terminal amino acids identical to those of the e. coli flavohemoglobin. furthermore, nod activity was produced by t ...19989724711
escherichia coli tol-pal mutants form outer membrane vesicles.mutations in the tol-pal genes induce pleiotropic effects such as release of periplasmic proteins into the extracellular medium and hypersensitivity to drugs and detergents. other outer membrane defective strains such as tolc, lpp, and rfa mutations are also altered in their outer membrane permeability. in this study, electron microscopy and western blot analyses were used to show that strains with mutations in each of the tol-pal genes formed outer membrane vesicles after growth in standard liq ...19989733690
the pir gene of erwinia chrysanthemi ec16 regulates hyperinduction of pectate lyase virulence genes in response to plant signals.the plant pathogenic bacterium erwinia chrysanthemi secretes pectate lyase proteins that are important virulence factors attacking the cell walls of plant hosts. bacterial production of these enzymes is induced by the substrate polypectate-na (napp) and further stimulated by the presence of plant extracts. the bacterial regulator responsible for induction by plant extracts was identified and purified by using a dna-binding assay with the promoter region of pele that encodes a major pectate lyase ...19989826648
colicin import into escherichia coli cells. 19989748429
mutation analysis of pobr and pcau, closely related transcriptional activators in acinetobacter.acinetobacter pobr and pcau are transcriptional activators that closely resemble each other in primary structure, dna-binding sites, metabolic modulators, and physiological function. pobr responds to the inducer-metabolite p-hydroxybenzoate and activates transcription of poba, the structural gene for the enzyme that converts p-hydroxybenzoate to protocatechuate. this compound, differing from p-hydroxybenzoate only in that it contains an additional oxygen atom, binds to pcau and thereby specifica ...19989748437
the virulence plasmid of yersinia, an antihost genome.the 70-kb virulence plasmid enables yersinia spp. (yersinia pestis, y. pseudotuberculosis, and y. enterocolitica) to survive and multiply in the lymphoid tissues of their host. it encodes the yop virulon, an integrated system allowing extracellular bacteria to disarm the cells involved in the immune response, to disrupt their communications, or even to induce their apoptosis by the injection of bacterial effector proteins. this system consists of the yop proteins and their dedicated type iii sec ...19989841674
hrpw of erwinia amylovora, a new harpin that contains a domain homologous to pectate lyases of a distinct class.harpins, such as hrpn of erwinia amylovora, are extracellular glycine-rich proteins that elicit the hypersensitive reaction (hr). we identified hrpw of e. amylovora, which encodes a protein similar to known harpins in that it is acidic, rich in glycine and serine, and lacks cysteine. a putative hrpl-dependent promoter was identified upstream of hrpw, and western blot analysis of hrpl mutants indicated that the production of hrpw is regulated by hrpl. hrpw is secreted via the hrp (type iii) pathw ...19989748455
development of a streptavidin-conjugated single-chain antibody that binds bacillus cereus spores.control of microorganisms such as bacillus cereus spores is critical to ensure the safety and a long shelf life of foods. a bifunctional single chain antibody has been developed for detection and binding of b. cereus t spores. the genes that encode b. cereus t spore single-chain antibody and streptavidin were connected for use in immunoassays and immobilization of the recombinant antibodies. a truncated streptavidin, which is smaller than but has biotin binding ability similar to that of strepta ...19989647821
engineering, expression and biochemical characterization of the hemoglobin domain of a erwinia chrysanthemi flavohemoprotein.an artificial hemoglobin-like domain has been constructed by engineering the gene coding for the multi-domain flavohemoprotein from the bacterium erwinia chrysanthemi. this domain was designed by molecular modelling, cloned and over-expressed in escherichia coli. the holo-protein was obtained in large quantities after extraction from inclusion bodies and refolding in presence of alkaline hemin. the purified 140-residue domain was studied and characterized to gain new insights into the biochemica ...19989654075
rpos (sigma-s) controls expression of rsma, a global regulator of secondary metabolites, harpin, and extracellular proteins in erwinia carotovora.rpos (sigma-s or sigma-38) controls a large array of genes that are expressed during stationary phase and under various stress conditions in escherichia coli and other bacteria. we document here that plant pathogenic and epiphytic erwinia species, such as e. amylovora; e. carotovora subsp. atroseptica, betavasculorum, and carotovora; e. chrysanthemi; e. herbicola; e. rhapontici; and e. stewartii, possess rpos genes and produce the alternate sigma factor. we show that rpos transcription in e. car ...19989658007
structural and biochemical analysis of the sheath of phormidium uncinatum.the sheath of the filamentous, gliding cyanobacterium phormidium uncinatum was studied by using light and electron microscopy. in thin sections and freeze fractures the sheath was found to be composed of helically arranged carbohydrate fibrils, 4 to 7 nm in diameter, which showed a substantial degree of crystallinity. as in all other examined motile cyanobacteria, the arrangement of the sheath fibrils correlates with the motion of the filaments during gliding motility; i.e., the fibrils formed a ...19989683490
cloning and expression of three new aazotobacter vinelandii genes closely related to a previously described gene family encoding mannuronan c-5-epimerases.the cloning and expression of a family of five modular-type mannuronan c-5-epimerase genes from azotobacter vinelandii (alge1 to -5) has previously been reported. the corresponding proteins catalyze the ca2+-dependent polymer-level epimerization of beta-d-mannuronic acid to alpha-l-guluronic acid (g) in the commercially important polysaccharide alginate. here we report the identification of three additional structurally similar genes, designated alge6, alge7, and algy. all three genes were seque ...19999864314
a novel beta-n-acetylglucosaminidase from streptomyces thermoviolaceus opc-520: gene cloning, expression, and assignment to family 3 of the glycosyl hydrolases.a beta-n-acetylglucosaminidase gene (naga) of streptomyces thermoviolaceus opc-520 was cloned in streptomyces lividans 66. the nucleotide sequence of the gene, which encodes naga, revealed an open reading frame of 1,896 bp, encoding a protein with an mr of 66, 329. the deduced primary structure of naga was confirmed by comparison with the n-terminal amino acid sequence of the cloned beta-n-acetylglucosaminidase expressed by s. lividans. the enzyme shares no sequence similarity with the classical ...19989687451
a cloned erwinia chrysanthemi hrp (type iii protein secretion) system functions in escherichia coli to deliver pseudomonas syringae avr signals to plant cells and to secrete avr proteins in culture.the hrp (type iii protein secretion) system is essential for the plant parasitic ability of pseudomonas syringae and most gram-negative bacterial plant pathogens. avrb and avrpto are two p. syringae proteins that have biological activity when produced via heterologous gene expression inside plant cells or when produced by hrp+ bacteria. avr-like proteins, presumably injected by the hrp system on bacterial contact with plant cells, appear to underlie pathogenic interactions, but none has been obs ...19989707625
negative regulation of hrp genes in pseudomonas syringae by hrpv.mutations in the five hrp and hrc genes in the hrpc operon of the phytopathogen pseudomonas syringae pv. syringae 61 have different effects on bacterial interactions with host and nonhost plants. the hrcc gene within the hrpc operon encodes an outer membrane component of the hrp secretion system that is conserved in all type iii protein secretion systems and is required for most pathogenic phenotypes and for secretion of the hrpz harpin to the bacterial milieu. the other four genes (in order), h ...19989721292
characterization of the hrpc and hrprs operons of pseudomonas syringae pathovars syringae, tomato, and glycinea and analysis of the ability of hrpf, hrpg, hrcc, hrpt, and hrpv mutants to elicit the hypersensitive response and disease in plants.the species pseudomonas syringae encompasses plant pathogens with differing host specificities and corresponding pathovar designations. p. syringae requires the hrp (type iii protein secretion) system, encoded by a 25-kb cluster of hrp and hrc genes, in order to elicit the hypersensitive response (hr) in nonhosts or to be pathogenic in hosts. dna sequence analysis of the hrpc and hrprs operons of p. syringae pv. syringae 61 (brown spot of beans), p. syringae pv. glycinea u1 (bacterial blight of ...19989721291
macromolecular assembly and secretion across the bacterial cell envelope: type ii protein secretion systems.a decade ago, pugsley and colleagues reported the existence of a large region of klebsiella dna, distinct from the klebsiella gene encoding pullulanase, which was necessary for secretion of this enzyme to the cell surface in escherichia coli (d'enfert et al., 1987a,b). the pul genes it contained proved to be the tip of an iceberg. the sequences reported before 1992 (d'enfert et al., 1987a,b; d'enfert & pugsley, 1989; pugsley & reyss, 1990; reyss & pugsley, 1990) included only one gene (puld) tha ...19989641973
construction and expression of a bifunctional single-chain antibody against bacillus cereus p6ores.the variable-region genes of monoclonal antibody against bacillus cereus spores were cloned from mouse hybridoma cells by reverse transcription-pcr. the heavy- and light-chain variable-region genes were connected by a 45-base linker dna to allow folding of the fusion protein into a functional tertiary structure. for detection of protein expression, a 10-amino-acid strep tag (biotin-like peptide) was attached to the c terminus of recombinant antibody as the reporter peptide. the single-chain anti ...19989647820
the evolutionary origin of the protein-translocating channel of chloroplastic envelope membranes: identification of a cyanobacterial homolog.the known envelope membrane proteins of the chloroplastic protein import apparatus lack sequence similarity to proteins of other eukaryotic or prokaryotic protein transport systems. however, we detected a putative homolog of the gene encoding toc75, the protein-translocating channel from the outer envelope membrane of pea chloroplasts, in the genome of the cyanobacterium synechocystis sp. pcc 6803. we investigated whether the low sequence identity of 21% reflects a structural and functional rela ...19999892711
characterization of the galu gene of streptococcus pneumoniae encoding a uridine diphosphoglucose pyrophosphorylase: a gene essential for capsular polysaccharide biosynthesis.the galu gene of streptococcus pneumoniae has been cloned and sequenced. escherichia coli cells harboring the recombinant plasmid pmmg2 (galu) overproduced a protein that has been shown to correspond to a uridine 5'-triphosphate:glucose-1-phosphate uridylyltransferase (uridine diphosphoglucose [udp-glc] pyrophosphorylase) responsible for the synthesis of udp-glc, a key compound in the biosynthesis of polysaccharides. a gene very similar to the s. pneumoniae galu has been found in a partial nucle ...19989841918
functional characterization of the erwinia chrysanthemi outs protein, an element of a type ii secretion system.secretion of pectate lyases and a cellulase occurs in erwinia chrysanthemi through a type ii secretion machinery, the out system. proper insertion of the secretin outd in the outer membrane requires the presence of outs. outs is an outer-membrane lipoprotein that interacts directly with outd. using ligand-blotting experiments, it has been shown that this interaction requires at least the 62 c-terminal amino acids of outd. when this domain was added to the c-terminal extremity of the secreted pec ...19989846757
pcr-based detection of the causal agent of watermark disease in willows (salix spp.)the watermark disease, caused by brenneria salicis (formerly erwinia salicis), is of significant concern wherever tree-forming willows are grown or occur naturally. the movement of infected, asymptomatic cuttings is a major cause of pathogen dispersal. a reliable and sensitive diagnostic procedure is necessary for the safe movement of willow planting material. we derived primers from the nucleotide sequence of the 16s rrna gene of b. salicis for the development of a pcr to detect this pathogen. ...19989758827
crystallization and preliminary x-ray analysis of a member of a new family of pectate lyases, peil from erwinia chrysanthemi.peil, a pectate lyase (e.c. 4.2.2.9) from e. chrysanthemi 3937 that is not homologous to the lyases with known structures, has been purified and crystallized by the hanging-drop method using a variety of organic solvents as precipitant. elongated lathes grown from poly(ethylene glycol) plus isopropanol belong to the space group p212121 with cell dimensions a = 55.5, b = 58.2, c = 16.4 a with a single molecule in the asymmetric unit. although complete data sets have been collected to 2.3 a resolu ...19989761915
campylobacter fetus surface layer proteins are transported by a type i secretion system.the virulence of campylobacter fetus, a bacterial pathogen of ungulates and humans, is mediated in part by the presence of a paracrystalline surface layer (s-layer) that confers serum resistance. the subunits of the s-layer are s-layer proteins (slps) that are secreted in the absence of an n-terminal signal sequence and attach to either type a or b c. fetus lipopolysaccharide in a serospecific manner. antigenic variation of multiple slps (encoded by sapa homologs) of type a strain 23d occurs by ...19989851986
use of l-asparaginase in childhood all.owing to the high efficacy of l-asparaginase in the treatment of acute lymphatic leukaemia the enzyme was introduced into the chemotherapy schedules for remission induction of this disease shortly after results of large-scale clinical trials had become available. since asparaginase monotherapy was associated with a high response rate but short remission duration, the enzyme is currently employed within the framework of combination chemotherapy schedules which achieve treatment response in about ...19989768345
characterization of the erwinia chrysanthemi expi-expr locus directing the synthesis of two n-acyl-homoserine lactone signal molecules.the plant pathogen erwinia chrysanthemi produces three acyl-homoserine lactones (acyl-hsls). one has been identified as n-(3-oxohexanoyl)-homoserine lactone (ohhl), and the two others were supposed to be n (hexanoyl)-homoserine lactone (hhl) and n-(decanoyl)-homoserine lactone (dhl). the genes for a quorum-sensing signal generator (expi) and a response regulator (expr) were cloned. these genes are convergently transcribed and display high similarity to the expi-expr genes of erwinia carotovora. ...19989781877
integration of the quorum-sensing system in the regulatory networks controlling virulence factor synthesis in erwinia chrysanthemi.the expi-expr locus drives a quorum-sensing system in the phytopathogenic bacterium, erwinia chrysanthemi. purified expr, an n-acyl homoserine lactone-responsive regulatory protein, binds to the promoter/operator region of the expi and expr genes. dnase i footprinting experiments showed that expr protects the regions between -66 and -40 from the p1 transcription initiation site of expl and between -54 and -18 from the expr transcription initiation site p1. the protected region overlaps the two e ...19989781878
identification and characterization of sira, an iron-regulated protein from staphylococcus aureus.the acquisition of iron by pathogenic bacteria is often a crucial step in establishing infection. to accomplish this, many bacteria, including staphylococcus aureus, produce low-molecular-weight iron-chelating siderophores. however, the secretion and transport of these molecules in gram-positive organisms are poorly understood. the sequence, organization, and regulation of genes involved in siderophore transport are conserved among gram-negative bacteria. we used this information to identify a p ...199910049373
processing of the pectate lyase peli by extracellular proteases of erwinia chrysanthemi 3937.erwinia chrysanthemi causes soft rot on various plants. the maceration of plant tissues is mainly due to the action of endopectate lyases. the e. chrysanthemi strain 3937 produces eight endopectate lyases (pela, pelb, pelc, peld, pele, peli, pell and pelz) that are secreted by the out pathway. the necrotic response elicited by the wild-type e. chrysanthemi strain on tobacco leaves is due to an extracellular protein secreted by the out machinery. purification of the active factor revealed that it ...19989781882
mechanism of osmotic activation of the quaternary ammonium compound transporter (qact) of lactobacillus plantarum.the accumulation of quaternary ammonium compounds in lactobacillus plantarum is mediated via a single transport system with a high affinity for glycine betaine (apparent km of 18 microm) and carnitine and a low affinity for proline (apparent km of 950 microm) and other analogues. mutants defective in the uptake of glycine betaine were generated by uv irradiation and selected on the basis of resistance to dehydroproline (dhp), a toxic proline analogue. three independent dhp-resistant mutants show ...19989791101
endochitinase is transported to the extracellular milieu by the eps-encoded general secretory pathway of vibrio cholerae.the chia gene of vibrio cholerae encodes a polypeptide which degrades chitin, a homopolymer of n-acetylglucosamine (glcnac) found in cell walls of fungi and in the integuments of insects and crustaceans. chia has a coding capacity corresponding to a polypeptide of 846 amino acids having a predicted molecular mass of 88.7 kda. a 52-bp region with promoter activity was found immediately upstream of the chia open reading frame. insertional inactivation of the chromosomal copy of the gene confirmed ...19989791107
detection of ralstonia solanacearum, which causes brown rot of potato, by fluorescent in situ hybridization with 23s rrna-targeted probes.during the past few years, ralstonia (pseudomonas) solanacearum race 3, biovar 2, was repeatedly found in potatoes in western europe. to detect this bacterium in potato tissue samples, we developed a method based on fluorescent in situ hybridization (fish). the nearly complete genes encoding 23s rrna of five r. solanacearum strains and one ralstonia pickettii strain were pcr amplified, sequenced, and analyzed by sequence alignment. this resulted in the construction of an unrooted tree and suppor ...19989797321
the pect repressor interacts with regulatory regions of pectate lyase genes in erwinia chrysanthemi.erwinia chrysanthemi is a broad host range phytopathogenic enterobacterium responsible for soft-rot disease of many plant species. the pect gene encodes a repressor that negatively regulates the expression of virulence factors, such as pectinases, motility or exopolysaccharide synthesis. the cloned pect gene was overexpressed using a phage t7 system. the purification of pect involved the use of a tsk-heparin column and delivered the pect protein that was purified to near homogeneity. the purifie ...19989804934
characterization of the dimerization domain in bglg, an rna-binding transcriptional antiterminator from escherichia coli.the escherichia coli transcriptional antiterminator protein bglg inhibits transcription termination of the bgl operon in response to the presence of beta-glucosides in the growth medium. bglg is an rna-binding protein that recognizes a specific sequence partially overlapping the two terminators within the bgl transcript. the activity of bglg is determined by its dimeric state which is modulated by reversible phosphorylation. thus, only the nonphosphorylated dimer binds to the rna target site and ...199910074067
corynebacterium glutamicum is equipped with four secondary carriers for compatible solutes: identification, sequencing, and characterization of the proline/ectoine uptake system, prop, and the ectoine/proline/glycine betaine carrier, ectp.gram-positive soil bacterium corynebacterium glutamicum uses the compatible solutes glycine betaine, proline, and ectoine for protection against hyperosmotic shock. osmoregulated glycine betaine carrier betp and proline permease putp have been previously characterized; we have identified and characterized two additional osmoregulated secondary transporters for compatible solutes in c. glutamicum, namely, the proline/ectoine carrier, prop, and the ectoine/glycine betaine/proline carrier, ectp. a ...19989811661
the staphylococcus aureus alternative sigma factor sigmab controls the environmental stress response but not starvation survival or pathogenicity in a mouse abscess model.the role of sigmab, an alternative sigma factor of staphylococcus aureus, has been characterized in response to environmental stress, starvation-survival and recovery, and pathogenicity. sigmab was mainly expressed during the stationary phase of growth and was repressed by 1 m sodium chloride. a sigb insertionally inactivated mutant was created. in stress resistance studies, sigmab was shown to be involved in recovery from heat shock at 54 degreesc and in acid and hydrogen peroxide resistance bu ...19989829915
ralstonia solanacearum pectin methylesterase is required for growth on methylated pectin but not for bacterial wilt virulenceralstonia (pseudomonas) solanacearum causes bacterial wilt, a serious disease of many crop plants. the pathogen produces several extracellular plant cell wall-degrading enzymes, including polygalacturonases (pgs) and pectin methylesterase (pme). pme removes methyl groups from pectin, thereby facilitating subsequent breakdown of this cell wall component by pgs, which are known bacterial wilt virulence factors. r. solanacearum pgs could not degrade 93% methylated pectin unless the substrate was fi ...19989835583
high-affinity transport of choline-o-sulfate and its use as a compatible solute in bacillus subtilis.we report here that the naturally occurring choline ester choline-o-sulfate serves as an effective compatible solute for bacillus subtilis, and we have identified a high-affinity atp-binding cassette (abc) transport system responsible for its uptake. the osmoprotective effect of this trimethylammonium compound closely matches that of the potent and widely employed osmoprotectant glycine betaine. growth experiments with a set of b. subtilis strains carrying defined mutations in the glycine betain ...19999925583
iron regulation and pathogenicity in erwinia chrysanthemi 3937: role of the fur repressor protein.low iron availability is a triggering signal for coordinated expression of the genes encoding pectate lyases pelb, pelc, peld, and pele, and chrysobactin iron transport functions, which are two main determinants of phytopathogenicity of the erwinia chrysanthemi strain 3937. the possible implication of the ferric uptake regulation (fur) protein in this process was investigated. the e. chrysanthemi fur gene was cloned by functional complementation of an escherichia coli fur mutant and sequenced. t ...19999926414
the minimal gene set member msra, encoding peptide methionine sulfoxide reductase, is a virulence determinant of the plant pathogen erwinia chrysanthemi.peptide methionine sulfoxide reductase (msra), which repairs oxidized proteins, is present in most living organisms, and the cognate structural gene belongs to the so-called minimum gene set [mushegian, a. r. & koonin, e. v., (1996) proc. natl. acad. sci. usa 93, 10268-10273]. in this work, we report that msra is required for full virulence of the plant pathogen erwinia chrysanthemi. the following differences were observed between the wild-type and a msra- mutant: (i) the msra- mutant was more s ...19999927663
genetic and transcriptional analyses of the vibrio cholerae mannose-sensitive hemagglutinin type 4 pilus gene locus.the mannose-sensitive hemagglutinin (msha) of the vibrio cholerae o1 el tor biotype is a member of the family of type 4 pili. type 4 pili are found on the surface of a variety of gram-negative bacteria and have demonstrated importance as host colonization factors, bacteriophage receptors, and mediators of dna transfer. the gene locus required for the assembly and secretion of the msha pilus has been localized to a 16.7-kb region of the v. cholerae chromosome. sixteen genes required for hemagglut ...19999973335
the structural and functional organization of h-ns-like proteins is evolutionarily conserved in gram-negative bacteria.the structural gene of the h-ns protein, a global regulator of bacterial metabolism, has been identified in the group of enterobacteria as well as in closely related bacteria, such as erwinia chrysanthemi and haemophilus influenzae. isolated outside these groups, the bph3 protein of bordetella pertussis exhibits a low amino acid conservation with h-ns, particularly in the n-terminal domain. to obtain information on the structure, function and/or evolution of h-ns, we searched for other h-ns-rela ...19999987132
origin of a chloroplast protein importer.during evolution, chloroplasts have relinquished the majority of their genes to the nucleus. the products of transferred genes are imported into the organelle with the help of an import machinery that is distributed across the inner and outer plastid membranes. the evolutionary origin of this machinery is puzzling because, in the putative predecessors, the cyanobacteria, the outer two membranes, the plasma membrane, and the lipopolysaccharide layer lack a functionally similar protein import syst ...19989861056
atypical genetic locus associated with constitutive production of enterocin b by enterococcus faecium bfe 900.a purified bacteriocin produced by enterococcus faecium bfe 900 isolated from black olives was shown by edman degradation and mass spectrometric analyses to be identical to enterocin b produced by e. faecium t136 from meat (p. casaus, t. nilsen, l. m. cintas, i. f. nes, p. e. hernández, and h. holo, microbiology 143:2287-2294, 1997). the structural gene was located on a 2.2-kb hindiii fragment and a 12.0-kb ecori chromosomal fragment. the genetic characteristics and production of entb by e. faec ...199910224016
prognostic significance of early response to a single dose of asparaginase in childhood acute lymphoblastic leukemia.the in vitro and in vivo efficacy of a single dose of asparaginase in children with newly diagnosed acute lymphoblastic leukemia and the correlation between in vitro and in vivo antileukemic response and long-term outcome were prospectively evaluated.199910029805
the myxococcus xanthus pilq (sgla) gene encodes a secretin homolog required for type iv pilus biogenesis, social motility, and development.the myxococcus xanthus sgla1 spontaneous mutation was originally isolated because it allowed dispersed cell growth in liquid yet retained the ability to form fruiting bodies. consequently, most of today's laboratory strains either contain the sgla1 mutation or were derived from strains that carry it. subsequent work showed that sgla was a gene for social gliding motility, a process which is mediated by type iv pili. here sgla is shown to map to the major pil cluster and to encode a 901-amino-aci ...19999864308
characterization of biosynthetic enzymes for ectoine as a compatible solute in a moderately halophilic eubacterium, halomonas elongata.1,4,5,6-tetrahydro-2-methyl-4-pyrimidinecarboxylic acid (ectoine) is an excellent osmoprotectant. the biosynthetic pathway of ectoine from aspartic beta-semialdehyde (asa), in halomonas elongata, was elucidated by purification and characterization of each enzyme involved. 2,4-diaminobutyrate (daba) aminotransferase catalyzed reversively the first step of the pathway, conversion of asa to daba by transamination with l-glutamate. this enzyme required pyridoxal 5'-phosphate and potassium ions for i ...19999864317
molecular and biochemical characterization of the protein template controlling biosynthesis of the lipopeptide lichenysin.lichenysins are surface-active lipopeptides with antibiotic properties produced nonribosomally by several strains of bacillus licheniformis. here, we report the cloning and sequencing of an entire 26.6-kb lichenysin biosynthesis operon from b. licheniformis atcc 10716. three large open reading frames coding for peptide synthetases, designated lica, licb (three modules each), and licc (one module), could be detected, followed by a gene, licte, coding for a thioesterase-like protein. the domain st ...19999864322
toward functional genomics in bacteria: analysis of gene expression in escherichia coli from a bacterial artificial chromosome library of bacillus cereus.as the study of microbes moves into the era of functional genomics, there is an increasing need for molecular tools for analysis of a wide diversity of microorganisms. currently, biological study of many prokaryotes of agricultural, medical, and fundamental scientific interest is limited by the lack of adequate genetic tools. we report the application of the bacterial artificial chromosome (bac) vector to prokaryotic biology as a powerful approach to address this need. we constructed a bac libra ...199910339608
assembly of xcpr in the cytoplasmic membrane is required for extracellular protein secretion in pseudomonas aeruginosa.a broad range of extracellular proteins secreted by pseudomonas aeruginosa use the type ii or general secretory pathway (gsp) to reach the medium. this pathway requires the expression of at least 12 xcp gene products. xcpr, a putative nucleotide-binding protein, is essential for the secretion process across the outer membrane even though the protein contains no hydrophobic sequence that could target or anchor it to the bacterial envelope. for a better understanding of the relationship between xc ...19999882649
regulation of hexuronate utilization in bacillus subtilis.we have identified a locus essential for galacturonate utilization in bacillus subtilis. genes homologous to escherichia coli and erwinia chrysanthemi glucuronate and galacturonate metabolic genes were found in a cluster consisting of 10 open reading frames (orfs) in the b. subtilis chromosome. a mutant of b. subtilis containing a replacement of the second and third orfs was unable to grow with galacturonate as its primary carbon source. galacturonate induced expression from a sigmaa-dependent p ...19999882655
the lysr homolog lrha promotes rpos degradation by modulating activity of the response regulator spre.synthesis of the ompf porin of escherichia coli is regulated in response to environmental and growth phase signals. in order to identify constituents of the various regulatory pathways involved in modulating ompf transcriptional expression, transposon insertion mutagenesis was performed and mutations that increased ompf'-lacz activity were identified as previously described. mutations mapping to a previously identified gene of unknown function, lrha, were obtained. we found that lrha, a lysr hom ...19999882671
the pect repressor coregulates synthesis of exopolysaccharides and virulence factors in erwinia chrysanthemi.erwinia chrysanthemi 3937 synthesizes an exopolysaccharide (eps) composed of rhamnose, galactose, and galacturonic acid. fourteen transcriptional fusions in genes required for eps synthesis, named eps, were obtained by tn5-b21 mutagenesis. eleven of them are clustered on the chromosome and are repressed by pect, a regulator of pectate lyase synthesis. in addition, expression of these fusions is repressed by the catabolite regulatory protein, crp, and induced in low osmolarity medium. the three o ...19999885192
thermoregulated expression and characterization of an nad(p)h-dependent 2-cyclohexen-1-one reductase in the plant pathogenic bacterium pseudomonas syringae pv. glycinea.the phytopathogenic bacterium pseudomonas syringae pv. glycinea pg4180.n9 causes bacterial blight of soybeans and preferably infects its host plant during periods of cold, humid weather conditions. to identify proteins differentially expressed at low temperatures, total cellular protein fractions derived from pg4180.n9 grown at 18 and 28 degreesc were separated by two-dimensional gel electrophoresis. of several proteins which appeared to be preferentially present at 18 degreesc, a 40-kda protein ...19999922244
the thioredoxin superfamily: redundancy, specificity, and gray-area genomics. 199910049365
roles of pseudomonas aeruginosa las and rhl quorum-sensing systems in control of twitching motility.pseudomonas aeruginosa is a ubiquitous environmental bacterium and an important human pathogen. the production of several virulence factors by p. aeruginosa is controlled through two quorum-sensing systems, las and rhl. we have obtained evidence that both the las and rhl quorum-sensing systems are also required for type 4 pilus-dependent twitching motility and infection by the pilus-specific phage d3112cts. mutants which lack the ability to synthesize pai-1, pai-2, or both autoinducers were sign ...199910049396
structure of a plant cell wall fragment complexed to pectate lyase c.the three-dimensional structure of a complex between the pectate lyase c (pelc) r218k mutant and a plant cell wall fragment has been determined by x-ray diffraction techniques to a resolution of 2.2 a and refined to a crystallographic r factor of 18.6%. the oligosaccharide substrate, alpha-d-galpa-([1-->4]-alpha-d-galpa)3-(1-->4)-d-galpa , is composed of five galacturonopyranose units (d-galpa) linked by alpha-(1-->4) glycosidic bonds. pelc is secreted by the plant pathogen erwinia chrysanthemi ...199910368179
molecular cloning, sequencing, and expression of a novel multidomain mannanase gene from thermoanaerobacterium polysaccharolyticum.the mana gene of thermoanaerobacterium polysaccharolyticum was cloned in escherichia coli. the open reading frame of mana is composed of 3,291 bases and codes for a preprotein of 1,097 amino acids with an estimated molecular mass of 119,627 da. the start codon is preceded by a strong putative ribosome binding site (taaggcggtg) and a putative -35 (ttcgc) and -10 (taaaat) promoter sequence. the mana of t. polysaccharolyticum is a modular protein. sequence comparison and biochemical analyses demons ...199910049399
the exopolygalacturonate lyase pelw and the oligogalacturonate lyase ogl, two cytoplasmic enzymes of pectin catabolism in erwinia chrysanthemi 3937.erwinia chrysanthemi 3937 secretes into the external medium several pectinolytic enzymes, among which are eight isoenzymes of the endo-cleaving pectate lyases: pela, pelb, pelc, peld, and pele (family 1); peli (family 4); pell (family 3); and pelz (family 5). in addition, one exo-cleaving pectate lyase, pelx (family 3), has been found in the periplasm of e. chrysanthemi. the e. chrysanthemi 3937 gene kdgc has been shown to exhibit a high degree of similarity to the genes pely of yersinia pseudot ...199910383957
secretion, localization, and antibacterial activity of tasa, a bacillus subtilis spore-associated protein.the synthesis and subcellular localization of the proteins that comprise the bacillus subtilis spore are under a variety of complex controls. to better understand these controls, we have identified and characterized a 31-kda sporulation protein, called tasa, which is secreted into the culture medium early in sporulation and is also incorporated into the spore. tasa synthesis begins approximately 30 min after the onset of sporulation and requires the sporulation transcription factor genes spo0h a ...199910049401
cloning of the escherichia coli endo-1,4-d-glucanase gene and identification of its product.a plasmid (pyp17) containing a genomic dna insert from escherichia coli k-12 that confers the ability to hydrolyze carboxymethylcellulose (cmc) was isolated from a genomic library constructed in the cosmid vector plafr3 in e. coli dh5alpha. a small 1.65-kb fragment, designated bcsc (pyp300), was sequenced and found to contain an orf of 1,104 bp encoding a protein of 368 amino acid residues, with a calculated molecular weight of 41,700 da. bcsc carries a typical prokaryotic signal peptide of 21 a ...199910102357
characterization of the exopolygalacturonate lyase pelx of erwinia chrysanthemi 3937.erwinia chrysanthemi 3937 secretes several pectinolytic enzymes, among which eight isoenzymes of pectate lyases with an endo-cleaving mode (pela, pelb, pelc, peld, pele, peli, pell, and pelz) have been identified. two exo-cleaving enzymes, the exopolygalacturonate lyase, pelx, and an exo-poly-alpha-d-galacturonosidase, pehx, have been previously identified in other e. chrysanthemi strains. using a genomic bank of a 3937 mutant with the major pel genes deleted, we cloned a pectinase gene identifi ...199910049400
disaccharides as a new class of nonaccumulated osmoprotectants for sinorhizobium meliloti.sucrose and ectoine (1,4,5,6-tetrahydro-2-methyl-4-pyrimidine carboxylic acid) are very unusual osmoprotectants for sinorhizobium meliloti because these compounds, unlike other bacterial osmoprotectants, do not accumulate as cytosolic osmolytes in salt-stressed s. meliloti cells. here, we show that, in fact, sucrose and ectoine belong to a new family of nonaccumulated sinorhizobial osmoprotectants which also comprises the following six disaccharides: trehalose, maltose, cellobiose, gentiobiose, ...199910103242
kdgrecc negatively regulates genes for pectinases, cellulase, protease, harpinecc, and a global rna regulator in erwinia carotovora subsp. carotovora.erwinia carotovora subsp. carotovora produces extracellular pectate lyase (pel), polygalacturonase (peh), cellulase (cel), and protease (prt). the concerted actions of these enzymes largely determine the virulence of this plant-pathogenic bacterium. e. carotovora subsp. carotovora also produces harpinecc, the elicitor of the hypersensitive reaction. we document here that kdgrecc (kdg, 2-keto-3-deoxygluconate; kdgr, general repressor of genes involved in pectin and galacturonate catabolism), a ho ...199910198003
azospirillum irakense produces a novel type of pectate lyase.the pela gene from the n2-fixing plant-associated bacterium azospirillum irakense, encoding a pectate lyase, was isolated by heterologous expression in escherichia coli. nucleotide sequence analysis of the region containing pela indicated an open reading frame of 1,296 bp, coding for a preprotein of 432 amino acids with a typical amino-terminal signal peptide of 24 amino acids. n-terminal amino acid sequencing confirmed the processing of the protein in e. coli at the signal peptidase cleavage si ...199910198006
cloning, expression, and nucleotide sequence of the pseudomonas aeruginosa 142 ohb genes coding for oxygenolytic ortho dehalogenation of halobenzoates.we have cloned and characterized novel oxygenolytic ortho-dehalogenation (ohb) genes from 2-chlorobenzoate (2-cba)- and 2,4-dichlorobenzoate (2,4-dcba)-degrading pseudomonas aeruginosa 142. among 3,700 escherichia coli recombinants, two clones, dh5alphaf'(pod22) and dh5alphaf'(pod33), converted 2-cba to catechol and 2,4-dcba and 2,5-dcba to 4-chlorocatechol. a subclone of pod33, plasmid pe43, containing the 3,687-bp minimized ohb dna region conferred to p. putida pb2440 the ability to grow on 2- ...199910224014
genetic diversity of african and worldwide strains of ralstonia solanacearum as determined by pcr-restriction fragment length polymorphism analysis of the hrp gene regionthe genetic diversity among a worldwide collection of 120 strains of ralstonia solanacearum was assessed by restriction fragment length polymorphism (rflp) analysis of amplified fragments from the hrp gene region. five amplified fragments appeared to be specific to r. solanacearum. fifteen different profiles were identified among the 120 bacterial strains, and a hierarchical cluster analysis distributed them into eight clusters. each cluster included strains belonging to a single biovar, except ...199910224018
the xanthomonas hrp type iii system secretes proteins from plant and mammalian bacterial pathogens.studies of essential pathogenicity determinants in gram-negative bacteria have revealed the conservation of type iii protein secretion systems that allow delivery of virulence factors into host cells from plant and animal pathogens. ten of 21 hrp proteins of the plant pathogen xanthomonas campestris pv. vesicatoria have been suggested to be part of a type iii machinery. here, we report the hrp-dependent secretion of two avirulence proteins, avrbs3 and avrrxv, by x. campestris pv. vesicatoria str ...199910430949
the avr (effector) proteins hrma (hoppsya) and avrpto are secreted in culture from pseudomonas syringae pathovars via the hrp (type iii) protein secretion system in a temperature- and ph-sensitive manner.we present here data showing that the avr proteins hrma and avrpto are secreted in culture via the native hrp pathways from pseudomonas syringae pathovars that produce these proteins. moreover, their secretion is strongly affected by the temperature and ph of the culture medium. both hrma and avrpto were secreted at their highest amounts when the temperature was between 18 and 22 degrees c and when the culture medium was ph 6.0. in contrast, temperature did not affect the secretion of hrpz. ph d ...199910438746
self-regulation of pir, a regulatory protein responsible for hyperinduction of pectate lyase in erwinia chrysanthemi ec16.previously, we have cloned and characterized the pir (plant inducible regulator) gene, which is responsible for hyperinduction of the synthesis of an isozyme of pectate lyase (ple) in erwinia chrysanthemi ec16 in the presence of potato extract and sodium polypectate (napp). the pir protein purified from escherichia coli overexpressing pir is able to bind to the promoter region of pir as a dimer. self-regulation of pir by its own translational product (pir) was suggested from the findings that pi ...199910226371
dsba is required for stable expression of outer membrane protein yscc and for efficient yop secretion in yersinia pestis.the role of the periplasmic disulfide oxidoreductase dsba in yop secretion was investigated in yersinia pestis. a y. pestis dsba mutant secreted reduced amounts of the v antigen and yops and expressed reduced amounts of the full-sized yscc protein. site-directed mutagenesis of the four cysteine residues present in the yscc protein resulted in defects similar to those found in the dsba mutant. these results suggest that yscc contains at least one disulfide bond that is essential for the function ...199910438793
growth of azospirillum irakense kbc1 on the aryl beta-glucoside salicin requires either sala or salb.the rhizosphere nitrogen-fixing bacterium azospirillum irakense kbc1 is able to grow on pectin and beta-glucosides such as cellobiose, arbutin, and salicin. two adjacent genes, sala and salb, conferring beta-glucosidase activity to escherichia coli, have been identified in a cosmid library of a. irakense dna. the sala and salb enzymes preferentially hydrolyzed aryl beta-glucosides. a delta(sala-salb) a. irakense mutant was not able to grow on salicin but could still utilize arbutin, cellobiose, ...199910321999
genetic and biochemical characterization of an exopolygalacturonase and a pectate lyase from yersinia enterocolitica.yersinia enterocolitica, an invasive foodborne human pathogen, degrades polypectate by producing two depolymerizing enzymes, pectate lyase (pl) and polygalacturonase (pg). the gene encoding the pg activity, designated pehy, was located in a 3-kb genomic fragment of y. enterocolitica atcc 49397. the complete nucleotide sequence of this 3-kb fragment was determined and an open reading frame consisting of 1803 bp was predicted to encode a pg protein with an estimated m(r) of 66 kda and pi of 6.3. t ...199910446714
direct interaction of the epsl and epsm proteins of the general secretion apparatus in vibrio cholerae.the general secretion pathway of gram-negative bacteria is responsible for extracellular secretion of a number of different proteins, including proteases and toxins. this pathway supports secretion of proteins across the cell envelope in two distinct steps, in which the second step, involving translocation through the outer membrane, is assisted by at least 13 different gene products. two of these components, the cytoplasmic membrane proteins epsl and epsm of vibrio cholerae, have been purified ...199910322014
enhancement of expression and apparent secretion of erwinia chrysanthemi endoglucanase (encoded by celz) in escherichia coli b.escherichia coli b has been engineered as a biocatalyst for the conversion of lignocellulose into ethanol. previous research has demonstrated that derivatives of e. coli b can produce high levels of erwinia chrysanthemi endoglucanase (encoded by celz) as a periplasmic product and that this enzyme can function with commercial fungal cellulase to increase ethanol production. in this study, we have demonstrated two methods that improve celz expression in e. coli b. initially, with a low-copy-number ...199910347024
cellobiose transport by mixed ruminal bacteria from a cow.the transport of cellobiose in mixed ruminal bacteria harvested from a holstein cow fed an italian ryegrass hay was determined in the presence of nojirimycin-1-sulfate, which almost inhibited cellobiase activity. the kinetic parameters of cellobiose uptake were 14 microm for the km and 10 nmol/min/mg of protein for the vmax. extracellular and cell-associated cellobiases were detected in the rumen, with both showing higher vmax values and lower affinities than those determined for cellobiose tran ...199910347044
purification and characterization of an extracellular protease from the fish pathogen yersinia ruckeri and effect of culture conditions on production.a novel protease, hydrolyzing azocasein, was identified, purified, and characterized from the culture supernatant of the fish pathogen yersinia ruckeri. exoprotease production was detected at the end of the exponential growth phase and was temperature dependent. activity was detected in peptone but not in casamino acid medium. its synthesis appeared to be under catabolite repression and ammonium control. the protease was purified in a simple two-step procedure involving ammonium sulfate precipit ...199910473403
regulation of hmp gene transcription in mycobacterium tuberculosis: effects of oxygen limitation and nitrosative and oxidative stress.the mycobacterium tuberculosis hmp gene encodes a protein which is homologous to flavohemoglobin in escherichia coli. northern blotting analysis demonstrated that hmp transcription increased when a microaerophilic culture became oxygen limited as it entered stationary phase at 20 days. there was a fivefold increase of the hmp transcripts during early stationary phase compared with the value which was observed in the exponential growth phase. this induction of hmp transcription was not due to cha ...199910348862
development and application of pathovar-specific monoclonal antibodies that recognize the lipopolysaccharide o antigen and the type iv fimbriae of xanthomonas hyacinthi.the objective of this study was to develop a specific immunological diagnostic assay for yellow disease in hyacinths, using monoclonal antibodies (mabs). mice were immunized with a crude cell wall preparation (shear fraction) from xanthomonas hyacinthi and with purified type iv fimbriae. hybridomas were screened for a positive reaction with x. hyacinthi cells or fimbriae and for a negative reaction with x. translucens pv. graminis or erwinia carotovora subsp. carotovora. nine mabs recognized fim ...199910473431
assembly of the type ii secretion machinery of erwinia chrysanthemi: direct interaction and associated conformational change between oute, the putative atp-binding component and the membrane protein outl.erwinia chrysanthemi secretes, by the type ii secretory pathway, a large number of enzymes, including cellulases and pectinases. this process requires the products of the out genes, which are widely conserved in gram-negative bacteria. the out proteins are thought to form a membrane-associated multiprotein complex. here, we investigated interaction between oute, the putative atp binding component, and outl, an inner membrane protein. we showed, by limited proteolysis, genetic suppression and the ...199910356336
modes of action of five different endopectate lyases from erwinia chrysanthemi 3937.five endopectate lyases from the phytopathogenic bacterium erwinia chrysanthemi, pela, pelb, peld, peli, and pell, were analyzed with respect to their modes of action on polymeric and oligomeric substrates (degree of polymerization, 2 to 8). on polygalacturonate, pelb showed higher reaction rates than peld, peli, and pela, whereas the reaction rates for pell were extremely low. the product progression during polygalacturonate cleavage showed a typical depolymerization profile for each enzyme and ...199910368144
molecular characterization of a protease secreted by erwinia amylovora.a protease with a molecular mass of 48 kda is secreted by the fire blight pathogen erwinia amylovora in minimal medium. we characterized this activity as a metalloprotease, since the enzyme was inhibited by edta and o -phenanthroline. a gene cluster was determined to encode four genes connected to protease expression, including a structural gene (prta) and three genes (prtd, prte, prtf) for secretion of the protease, which are transcribed in the same direction. the organization of the protease g ...199910373365
mutational analysis of the recj exonuclease of escherichia coli: identification of phosphoesterase motifs.the recj gene, identified in escherichia coli, encodes a mg(+2)-dependent 5'-to-3' exonuclease with high specificity for single-strand dna. genetic and biochemical experiments implicate recj exonuclease in homologous recombination, base excision, and methyl-directed mismatch repair. genes encoding proteins with strong similarities to recj have been found in every eubacterial genome sequenced to date, with the exception of mycoplasma and mycobacterium tuberculosis. multiple genes encoding protein ...199910498723
elements involved in catabolite repression and substrate induction of the lactose operon in lactobacillus casei.in lactobacillus casei atcc 393, the chromosomally encoded lactose operon, lactegf, encodes an antiterminator protein (lact), lactose-specific phosphoenolpyruvate-dependent phosphotransferase system (pts) elements (lace and lacf), and a phospho-beta-galactosidase. lact, lace, and lacf mutant strains were constructed by double crossover. the lact strain displayed constitutive termination at a ribonucleic antiterminator (rat) site, whereas lace and lacf mutants showed an inducer-independent antite ...199910383959
opening the iron box: transcriptional metalloregulation by the fur protein. 199910515908
ferric enterochelin transport in yersinia enterocolitica: molecular and evolutionary aspects.yersinia enterocolitica is well equipped for siderophore piracy, encompassing the utilization of siderophores such as ferrioxamine, ferrichrome, and ferrienterochelin. in this study, we report on the molecular and functional characterization of the yersinia fep-fes gene cluster orthologous to the escherichia coli ferrienterochelin transport genes (fepa, fepdgc, and fepb) and the esterase gene fes. in vitro transcription-translation analysis identified polypeptides of 30 and 35 kda encoded by fep ...199910515929
the mdoc gene of escherichia coli encodes a membrane protein that is required for succinylation of osmoregulated periplasmic glucans.osmoregulated periplasmic glucans (opgs) of escherichia coli are anionic oligosaccharides that accumulate in the periplasmic space in response to low osmolarity of the medium. their anionic character is provided by the substitution of the glucosidic backbone by phosphoglycerol originating from the membrane phospholipids and by succinyl residues from unknown origin. a phosphoglycerol-transferase-deficient mdob mutant was subjected to tn5 transposon mutagenesis, and putative mutant clones were scr ...199910368134
osmosensing by bacteria: signals and membrane-based sensors.bacteria can survive dramatic osmotic shifts. osmoregulatory responses mitigate the passive adjustments in cell structure and the growth inhibition that may ensue. the levels of certain cytoplasmic solutes rise and fall in response to increases and decreases, respectively, in extracellular osmolality. certain organic compounds are favored over ions as osmoregulatory solutes, although k+ fluxes are intrinsic to the osmoregulatory response for at least some organisms. osmosensors must undergo tran ...199910066837
identification of the tlidef abc transporter specific for lipase in pseudomonas fluorescens sik w1.pseudomonas fluorescens, a gram-negative psychrotrophic bacterium, secretes a thermostable lipase into the extracellular medium. in our previous study, the lipase of p. fluorescens sik w1 was cloned and expressed in escherichia coli, but it accumulated as inactive inclusion bodies. amino acid sequence analysis of the lipase revealed a potential c-terminal targeting sequence recognized by the atp-binding cassette (abc) transporter. the genetic loci around the lipase gene were searched, and a secr ...199910074078
the mxi-spa type iii secretory pathway of shigella flexneri requires an outer membrane lipoprotein, mxim, for invasin translocation.invasion of epithelial cells by shigella flexneri is mediated by a set of translocated bacterial invasins, the ipa proteins, and its dedicated type iii secretion system, called mxi-spa. we show here that mxim, part of the mxi-spa locus in the s. flexneri virulence plasmid, encodes an indispensable type iii secretion apparatus component, required for both ipa translocation and tissue culture cell invasion. we demonstrated that mature mxim, first identified as a putative lipoprotein, is lipidated ...199910085046
badr, a new marr family member, regulates anaerobic benzoate degradation by rhodopseudomonas palustris in concert with aadr, an fnr family member.a cluster of genes for the anaerobic degradation of benzoate has been described for the phototrophic bacterium rhodopseudomonas palustris. here we provide an initial analysis of the regulation of anaerobic benzoate degradation by examining the contributions of two regulators: a new regulator, badr, encoded by the benzoate degradation gene cluster, and a previously described regulator, aadr, whose gene lies outside the cluster. strains with single mutations in either badr or aadr grew slowly on b ...199910094687
cellular locations of pseudomonas syringae pv. syringae hrcc and hrcj proteins, required for harpin secretion via the type iii pathway.the complete hrp-hrc-hrma cluster of pseudomonas syringae pv. syringae 61 encodes 28 polypeptides. a saprophytic bacterium carrying this cluster is capable of secreting hrpz-a harpin encoded by hrpz-in an hrp-dependent manner, which suggests that this cluster contains sufficient components to assemble functional type iii secretion machinery. sequence data show that hrcj and hrcc are putative outer membrane proteins, and nonpolar mutagenesis demonstrates they are all required for hrpz secretion. ...199910094714
the glucuronic acid utilization gene cluster from bacillus stearothermophilus t-6.a lambda-embl3 genomic library of bacillus stearothermophilus t-6 was screened for hemicellulolytic activities, and five independent clones exhibiting beta-xylosidase activity were isolated. the clones overlap each other and together represent a 23.5-kb chromosomal segment. the segment contains a cluster of xylan utilization genes, which are organized in at least three transcriptional units. these include the gene for the extracellular xylanase, xylanase t-6; part of an operon coding for an intr ...199910368143
membrane association and multimerization of secreton component pulc.the pulc component of the klebsiella oxytoca pullulanase secretion machinery (the secreton) was found by subcellular fractionation to be associated with both the cytoplasmic (inner) and outer membranes. association with the outer membrane was independent of other secreton components, including the outer membrane protein puld (secretin). the association of pulc with the inner membrane is mediated by the signal anchor sequence located close to its n terminus. these results suggest that pulc forms ...199910383968
structure-function analysis of xcpp, a component involved in general secretory pathway-dependent protein secretion in pseudomonas aeruginosa.the general secretory pathway of pseudomonas aeruginosa is required for the transport of signal peptide-containing exoproteins across the cell envelope. after completion of the sec-dependent translocation of exoproteins across the inner membrane and cleavage of the signal peptide, the xcp machinery mediates translocation across the outer membrane. this machinery consists of 12 components, of which xcpq (gspd) is the sole outer membrane protein. xcpq forms a multimeric ring-shaped structure, with ...199910383969
secretin puld: association with pilot puls, structure, and ion-conducting channel formation.the outer membrane protein puld (secretin) of klebsiella oxytoca is required for transport of pullulanase across this membrane. we have purified a multimeric puld complex from an escherichia coli strain expressing all the proteins involved in pullulanase secretion. the outer membrane-anchored lipoprotein puls was found to copurify with puld. the molar ratio of the two proteins is close to 1:1, and the size of the complex is approximately 1 mda. scanning transmission electron and cryo-electron mi ...199910393967
acquisition, transport, and storage of iron by pathogenic fungi.iron is required by most living systems. a great variety of means of acquisition, avenues of uptake, and methods of storage are used by pathogenic fungi to ensure a supply of the essential metal. solubilization of insoluble iron polymers is the first step in iron assimilation. the two methods most commonly used by microorganisms for solubilization of iron are reduction and chelation. reduction of ferric iron to ferrous iron by enzymatic or nonenzymatic means is a common mechanism among pathogeni ...199910398672
functional analysis of the carbohydrate-binding domains of erwinia chrysanthemi cel5 (endoglucanase z) and an escherichia coli putative chitinase.the cel5 cellulase (formerly known as endoglucanase z) from erwinia chrysanthemi is a multidomain enzyme consisting of a catalytic domain, a linker region, and a cellulose binding domain (cbd). a three-dimensional structure of the cbd(cel5) has previously been obtained by nuclear magnetic resonance. in order to define the role of individual residues in cellulose binding, site-directed mutagenesis was performed. the role of three aromatic residues (trp18, trp43, and tyr44) in cellulose binding wa ...199910419961
extracellular polysaccharide of erwinia chrysanthemi cu643.erwinia chrysanthemi are gram-negative bacterial phytopathogens causing soft rots in a number of plants. the structure of the extracellular polysaccharide (eps) produced by e. chrysanthemi strain cu643, pathogenic to philodendron, has been determined using a combination of chemical and physical techniques including methylation analysis, high- and low-pressure gel-filtration and anion-exchange chromatography, high-ph anion-exchange chromatography, partial acid hydrolysis, mass spectrometry, and 1 ...199910420593
cloning and sequencing of a high-alkaline pectate lyase gene from an alkaliphilic bacillus isolate.alkaliphilic bacillus sp. strain ksm-p103 was found to exoproduce a high-alkaline pectate lyase (pectate transeliminase, ec 4.2.2.2). the gene for this enzyme from the alkaliphile was cloned and sequenced for the first time. the structural gene contained a 1,038-bp open reading frame encoding 345 amino acids. the deduced amino acid sequence of the mature enzyme (302 amino acids, 33,312 da), designated pel-103, showed very low similarity to those of known pectate lyases with 28-36% identity: the ...199910427684
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