Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Acinetobacter”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,549 records · Page 86Linked to original sources
PubMed1995

catM encodes a LysR-type transcriptional activator regulating catechol degradation in Acinetobacter calcoaceticus.

On the basis of the constitutive phenotypes of two catM mutants of Acinetobacter calcoaceticus, the CatM protein was proposed to repress expression of two different loci involved in catechol degradation, catA and catBCIJFD (E. Neidle, C. Hartnett, and L. N. Ornston, J. Bacteriol. 171:5410-5421, 1989). In spite of its proposed negative role as a repressor, CatM is similar in amino acid sequence to positive transcriptional activators of the LysR family. Investigating this anomaly, we found that insertional inactivation of catM did not cause the phenotype expected for the disruption of a repressor-encoding gene: in an interposon-generated catM mutant, no cat genes were expressed constitutively, but rather catA and catB were still inducible by muconate. Moreover, this catM mutant grew poorly on benzoate, a process requiring the expression of all cat genes. The inducibility of the cat genes in this catM mutant was completely eliminated by a 3.5-kbp deletion 10 kbp upstream of catM. In this double mutant, catM in trans restored muconate inducibility to both catA and catB. These results suggested the presence of an additional regulatory locus controlling cat gene expression. The ability of CatM to function as an activator was also suggested by these results. In support of this hypothesis, in vivo methylation protection assays showed that CatM protects two guanines in a dyad 65 nucleotides upstream of the catB transcriptional start site, in a location and pattern typical of LysR-type transcriptional activators. Gel mobility shift assays indicated that CatM also binds to a region upstream of catA. DNA sequence analysis revealed a nucleotide near the 3' end of catM not present in the published sequence. Translation of the corrected sequence resulted in the deduced CatM protein being 52 residues longer than previously reported. The size, amino acid sequence, and mode of action of CatM now appear similar to, and typical of, what has been found for transcriptional activators in the LysR family. Analysis of one of the constitutive alleles of catM previously thought to encode a dysfunctional repressor indicated instead that it encodes an inducer-independent transcriptional activator.

Acinetobacter calcoaceticus↗
PubMed1995

Unusual ancestry of dehydratases associated with quinate catabolism in Acinetobacter calcoaceticus.

Catabolism of quinate to protocatechuate requires the consecutive action of quinate dehydrogenase (QuiA), dehydroquinate dehydratase (QuiB), and dehydroshikimate dehyratase (QuiC), Genes for catabolism of protocatechuate are encoded by the pca operon in the Acinetobacter calcoaceticus chromosome. Observations reported here demonstrate that A. calcoaceticus qui genes are clustered in the order quiBCXA directly downstream from the pca operon. Sequence comparisons indicate that quiX encodes a porin, but the specific function of this protein has not been clearly established. Properties of mutants created by insertion of omega elements show that quiBC is expressed as part of a single transcript, but there is also an independent transcriptional initiation site directly upstream of quiA. The deduced amino acid sequence of QuiC does not resemble any other known sequence. A. calcoaceticus QuiB is most directly related to a family of enzymes with identical catalytic activity and biosynthetic AroD function in coliform bacteria. Evolution of A. calcoaceticus quiB appears to have been accompanied by fusion of a leader sequence for transport of the encoded protein into the inner membrane, and the location of reactions catalyzed by the mature enzyme may account for the failure of A. calcoaceticus aroD to achieve effective complementation of null mutations in quiB. Analysis of a genetic site where a DNA segment encoding a leader sequence was transposed adds to evidence suggesting horizontal transfer of nucleotide sequences within genes during evolution.

Acinetobacter calcoaceticus↗
PubMed1995

Distance between alleles as a determinant of linkage in natural transformation of Acinetobacter calcoaceticus.

Cotransformation frequencies of 16, 39, 51, and 60% were observed when donor alleles were separated by distances of 9.2, 7.4, 6.3, and 5.1 kb, respectively, in donor Acinetobacter calcoaceticus DNA. A different and unexpected pattern was observed when the distance between recipient alleles was reduced from 9.2 to 5.1 kb. Ligation of unlinked chromosomal DNA fragments allowed them to be linked genetically through natural transformation.

Acinetobacter calcoaceticus↗
PubMed1995

Spontaneous mutations in pcaH and -G, structural genes for protocatechuate 3,4-dioxygenase in Acinetobacter calcoaceticus.

Bacteria containing spontaneous null mutations in pcaH and -G, structural genes for protocatechuate 3,4-dioxygenase, were selected by exposure of an Acinetobacter calcoaceticus strain to physiological conditions in which expression of the genes prevents growth. The parental bacterial strain exhibits high competence for natural transformation, and this procedure was used to characterize 94 independently isolated spontaneous mutations. Four of the mutations were caused by integration of a newly identified insertion sequence, IS1236. Many (22 of 94) of the mutations were lengthy deletions, the largest of which appeared to eliminate at least 17 kb of DNA containing most of the pca-qui-pob supraoperonic gene cluster. DNA sequence determination revealed that the endpoints of four smaller deletions (74 to 440 bp in length) contained DNA sequence repetitions aligned imprecisely with the sites of mutation. Analysis of direct and inverted DNA sequence repetitions associated with the sites of mutation suggested the existence of DNA slippage structures that make unhybridized nucleotides particularly susceptible to mutation.

Acinetobacter calcoaceticus↗
PubMed1996

Physiological factors affecting production of extracellular lipase (LipA) in Acinetobacter calcoaceticus BD413: fatty acid repression of lipA expression and degradation of LipA.

The extracellular lipase (LipA) produced by Acinetobacter calcoaceticus BD413 is required for growth of the organism on triolein, since mutant strains that lack an active lipase fail to grow with triolein as the sole carbon source. Surprisingly, extracellular lipase activity and expression of the structural lipase gene (lipA), the latter measured through lacZ as a transcriptional reporter, are extremely low in triolein cultures of LipA+ strains. The explanation for this interesting paradox lies in the effect of fatty acids on the expression of lipA. We found that long-chain fatty acids, especially, strongly repress the expression of lipA, thereby negatively influencing the production of lipase. We propose the involvement of a fatty acyl-responsive DNA-binding protein in regulation of expression of the A. calcoaceticus lipBA operon. The potential biological significance of the observed physiological competition between expression and repression of lipA in the triolein medium is discussed. Activity of the extracellular lipase is also negatively affected by proteolytic degradation, as shown in in vitro stability experiments and by Western blotting (immunoblotting) of concentrated supernatants of stationary-phase cultures. In fact, the relatively high levels of extracellular lipase produced in the early stationary phase in media which contain hexadecane are due only to enhanced stability of the extracellular enzyme under those conditions. The rapid extracellular degradation of LipA of A. calcoaceticus BD413 by an endogenous protease is remarkable and suggests that proteolytic degradation of the enzyme is another important factor in regulating the level of active extracellular lipase.

Acinetobacter calcoaceticus↗
PubMed1996

Novel nuclear magnetic resonance spectroscopy methods demonstrate preferential carbon source utilization by Acinetobacter calcoaceticus.

Novel nuclear magnetic resonance spectroscopy techniques, designated metabolic observation, were used to study aromatic compound degradation by the soil bacterium Acinetobacter calcoaceticus. Bacteria which had been rendered spectroscopically invisible by growth with deuterated (2H) medium were used to inoculate cultures in which natural-abundance 1H hydrogen isotopes were provided solely by aromatic carbon sources in an otherwise 2H medium. Samples taken during the incubation of these cultures were analyzed by proton nuclear magnetic resonance spectroscopy, and proton signals were correlated with the corresponding aromatic compounds or their metabolic descendants. This approach allowed the identification and quantitation of metabolites which accumulated during growth. This in vivo metabolic monitoring facilitated studies of catabolism in the presence of multiple carbon sources, a topic about which relatively little is known. A. calcoaceticus initiates aromatic compound dissimilation by forming catechol or protocatechuate from a variety of substrates. Degradation proceeds via the beta-ketoadipate pathway, comprising two discrete branches that convert catechol or protocatechuate to tricarboxylic acid cycle intermediates. As shown below, when provided with several carbon sources simultaneously, all degraded via the beta-ketoadipate pathway, A. calcoaceticus preferentially degraded specific compounds. For example, benzoate, degraded via the catechol branch, was consumed in preference to p-hydroxybenzoate, degraded via the protocatechuate branch, when both compounds were present. To determine if this preference were governed by metabolites unique to catechol degradation, pathway mutants were constructed. Studies of these mutants indicated that the product of catechol ring cleavage, cis,cis-muconate, inhibited the utilization of p-hydroxybenzoate in the presence of benzoate. The accumulation of high levels of cis,cis-muconate also appeared to be toxic to the cells.

Acinetobacter calcoaceticus↗
PubMed1997

Identification and characterization of xcpR encoding a subunit of the general secretory pathway necessary for dodecane degradation in Acinetobacter calcoaceticus ADP1.

A mutant of Acinetobacter calcoaceticus ADP1 unable to grow on alkanes was complemented for growth on hexadecane with a DNA fragment encoding a protein with homology to XcpR, a subunit of the general secretion pathway for exoproteins in Pseudomonas aeruginosa. Insertional inactivation of xcpR in A. calcoaceticus ADP1 by transcriptional fusion to lacZ abolishes secretion of lipase and esterase and leads to lack of growth on dodecane and slower growth on hexadecane. We, therefore, propose the participation of a secreted protein in alkane degradation.

Acinetobacter calcoaceticus↗
PubMed1997

Cloning and characterization of two catA genes in Acinetobacter lwoffii K24.

Two novel type I catechol 1,2-dioxygenases inducible on aniline media were isolated from Acinetobacter lwoffii K24. Although the two purified enzymes, CD I1 and CD I2, had similar intradiol cleavage activities, they showed different substrate specificities for catechol analogs, physicochemical properties, and amino acid sequences. Two catA genes, catA1 and catA2, encoding by CD I1 and CD I2, respectively, were isolated from the A. lwoffii K24 genomic library by using colony hybridization and PCR. Two DNA fragments containing the catA1 and catA2 genes were located on separate regions of the chromosome. They contained open reading frames encoding 33.4- and 30.4-kDa proteins. The amino acid sequences of the two proteins matched well with previously determined sequences. Interestingly, further analysis of the two DNA fragments revealed the locations of the catB and catC genes as well. Moreover, the DNA fragment containing catA1 had a cluster of genes in the order catB1-catC1-catA1 while the catB2-catA2-catC2 arrangement was found in the catA2 DNA fragment. These results may provide an explanation of the different substrate specificities and physicochemical properties of CD I1 and CD I2.

Acinetobacter↗
PubMed1997

mucK, a gene in Acinetobacter calcoaceticus ADP1 (BD413), encodes the ability to grow on exogenous cis,cis-muconate as the sole carbon source.

Benzyl alcohol, benzaldehyde, benzoate, and anthranilate are metabolized via catechol, cis,cis-muconate, and the beta-ketoadipate pathway in Acinetobacter calcoaceticus ADP1 (BD413). Mutant strain ISA25 with a deletion spanning catBCIJF and unable to metabolize muconate further will not grow in the presence of an aromatic precursor of muconate. Growth on fumarate as the sole carbon source with added benzyl alcohol or benzaldehyde selected spontaneous mutants of ISA25. After repair of the cat deletion by natural transformation with linearized plasmid pPAN4 (catBCIJF) 10 mutants were unable to grow on benzoate of cis,cis-muconate but could still grow on anthranilate. Transformation with wild-type chromosomal DNA demonstrated the presence of two unlinked mutations in each strain, one in the benABCD region, encoding the conversion of benzoate to catechol, and the other in a gene determining the ability to grow on exogenous cis,cis-muconate. The wild-type gene, named mucK, was cloned into pUC18, and its nucleotide sequence was determined. It encodes a 413-residue protein of M(r) = 45,252 which is a member of a superfamily of membrane transport proteins and which is within a subgroup involved in the uptake of organic acids. Five of the mutant alleles were cloned, and the mutations were determined by nucleotide sequencing. All the mutations were in the mucK coding region and consisted of three deletions, one duplication, and a substitution. Insertional inactivation of mucK resulted in the loss of the ability to utilize exogenous muconate. The location of mucK on the chromosome appeared to be unique for genes associated with the benzoate branch of the beta-ketoadipate pathway in being close to the pca-qui-pob gene cluster (for p-hydroxybenzoate utilization) and distant from the functionally related ben-cat cluster. Downstream of mucK and transcribed in the same direction is an open reading frame encoding a protein of 570 residues (M(r) = 63,002) which shows considerable homology with a mammalian electron transport protein; its insertional inactivation had no detectable phenotypic effect.

Acinetobacter calcoaceticus↗
PubMed1997

benK encodes a hydrophobic permease-like protein involved in benzoate degradation by Acinetobacter sp. strain ADP1.

The chromosomal benK gene was identified within a supraoperonic gene cluster involved in benzoate degradation by Acinetobacter sp. strain ADP1, and benK was expressed in response to a benzoate metabolite, cis,cis-muconate. The disruption of benK reduced benzoate uptake and impaired the use of benzoate or benzaldehyde as the carbon source. BenK was homologous to several aromatic compound transporters.

Acinetobacter↗
PubMed1997

Dihydroxyacetone synthase from a methanol-utilizing carboxydobacterium, Acinetobacter sp. strain JC1 DSM 3803.

Acinetobacter sp. strain JC1 DSM 3803, a carboxydobacterium, grown on methanol was found to show dihydroxyacetone synthase, dihydroxyacetone kinase, and ribulose 1,5-bisphosphate carboxylase, but no hydroxypyruvate reductase and very low hexulose 6-phosphate synthase, activities. The dihydroxyacetone synthase was found to be expressed earlier than the ribulose 1,5-bisphosphate carboxylase. The dihydroxyacetone synthase was purified 19-fold in eight steps to homogeneity, with a yield of 9%. The final specific activity of the purified enzyme was 1.12 micromol of NADH oxidized per min per mg of protein. The molecular weight of the native enzyme was determined to be 140,000. Sodium dodecyl sulfate-gel electrophoresis revealed a subunit of molecular weight 73,000. The optimum temperature and pH were 30 degrees C and 7.0, respectively. The enzyme was inactivated very rapidly at 70 degrees C. The enzyme required Mg2+ and thiamine pyrophosphate for maximal activity. Xylulose 5-phosphate was found to be the best substrate when formaldehyde was used as a glycoaldehyde acceptor. Erythrose 4-phosphate, glycolaldehyde, and formaldehyde were found to act as excellent substrates when xylulose 5-phosphate was used as a glycoaldehyde donor. The Kms for formaldehyde and xylulose 5-phosphate were 1.86 mM and 33.3 microM, respectively. The enzyme produced dihydroxyacetone from formaldehyde and xylulose 5-phosphate. The enzyme was found to be expressed only in cells grown on methanol and shared no immunological properties with the yeast dihydroxyacetone synthase.

Acinetobacter↗
PubMed1997

Expression, inducer spectrum, domain structure, and function of MopR, the regulator of phenol degradation in Acinetobacter calcoaceticus NCIB8250.

Degradation of phenol by Acinetobacter calcoaceticus NCIB8250 involves (sigma54-dependent expression of a multicomponent phenol hydroxylase and catechol 1,2-dioxygenase encoded by the mop operon. Complementation of a new mutant deficient in phenol utilization yielded the regulatory locus mopR. It is located in divergent orientation next to the mop operon. MopR is constitutively expressed at a low level from a sigma70-type promoter and belongs to the NtrC family of regulators. The amino acid sequence is similar to that of XylR regulating xylene degradation and to that of DmpR regulating dimethylphenol degradation in Pseudomonas spp. However, it shows a different effector profile for substituted phenols than DmpR. MopR activates phenol hydroxylase expression in the presence of phenol in Escherichia coli, indicating that it binds the effector. The phenol binding A domains of MopR and DmpR have fewer identical residues than the A domains of DmpR and XylR, despite the fact that XylR recognizes different effectors. This suggests that sequence conservation in the A domain does not reflect the potential to bind the respective effectors. Overexpression of the MopR A domain in the presence of wild-type MopR causes loss of mop inducibility by phenol, establishing its negative transdominance over MopR. Deletion of 110 residues from the N terminus did not affect transdominance of the truncated domain, whereas deletion of 150 residues abolished it completely. This result establishes the distinction of two subdomains, A(N) and A(C), which together constitute the A domain. The C-terminal portion of the A domain, A(C), shows considerable affinity for the C domain, even in the presence of the trigger phenol.

Acinetobacter calcoaceticus↗
PubMed1997

Isolation of mutants of Acinetobacter calcoaceticus deficient in wax ester synthesis and complementation of one mutation with a gene encoding a fatty acyl coenzyme A reductase.

Acinetobacter calcoaceticus BD413 accumulates wax esters and triacylglycerol under conditions of mineral nutrient limitation. Nitrosoguanidine-induced mutants of strain BD413 were isolated that failed to accumulate wax esters under nitrogen-limited growth conditions. One of the mutants, Wow15 (without wax), accumulated wax when grown in the presence of cis-11-hexadecenal and hexadecanol but not hexadecane or hexadecanoic acid. This suggested that the mutation may have inactivated a gene encoding either an acyl-acyl carrier protein or acyl-coenzyme A (CoA) reductase. The Wow15 mutant was complemented with a cosmid genomic library prepared from wild-type A. calcoaceticus BD413. The complementary region was localized to a single gene (acr1) encoding a protein of 32,468 Da that is 44% identical over a region of 264 amino acids to a product of unknown function encoded by an open reading frame associated with mycolic acid synthesis in Mycobacterium tuberculosis H37Ra. Extracts of Escherichia coli cells expressing the acr1 gene catalyzed the reduction of acyl-CoA to the corresponding fatty aldehyde, indicating that the gene encodes a novel fatty acyl-CoA reductase.

Acinetobacter calcoaceticus↗
PubMed1998

Expression of alkane hydroxylase from Acinetobacter sp. Strain ADP1 is induced by a broad range of n-alkanes and requires the transcriptional activator AlkR.

In Acinetobacter sp. strain ADP1, alkane degradation depends on at least five essential genes. rubAB and xcpR are constitutively transcribed. Here we describe inducible transcription of alkM, which strictly depends on the presence of the transcriptional activator AlkR. alkR itself is expressed at a low level, while a chromosomally located alkM::lacZ fusion is inducible by middle-chain-length alkanes from heptane to undecane, which do not support growth of ADP1, and by long-chain-length alkanes from dodecane to octadecane, which are used as sources of carbon and energy. The putative AlkM substrate 1-dodecene is also an effective inducer. Products of alkane hydroxylase activity like 1-dodecanol prevent induction of alkM expression. alkM is expressed only in stationary phase, suggesting its dependence on at least one other regulatory mechanism.

Acinetobacter↗
PubMed1998

Identification and characterization of a novel competence gene, comC, required for DNA binding and uptake in Acinetobacter sp. strain BD413.

A gene (comC) essential for natural transformation was identified in Acinetobacter sp. strain BD413. ComC has a typical leader sequence and is similar to different type IV pilus assembly factors. A comC mutant (T308) is not able to bind or take up DNA but exhibits a piliation phenotype indistinguishable from the transformation wild type as revealed by electron microscopy.

Acinetobacter↗
PubMed1998

Regulation of benzoate degradation in Acinetobacter sp. strain ADP1 by BenM, a LysR-type transcriptional activator.

In Acinetobacter sp. strain ADP1, benzoate degradation requires the ben genes for converting benzoate to catechol and the cat genes for degrading catechol. Here we describe a novel transcriptional activator, BenM, that regulates the chromosomal ben and cat genes. BenM is homologous to CatM, a LysR-type transcriptional activator of the cat genes. Unusual regulatory features of this system include the abilities of both BenM and CatM to recognize the same inducer, cis,cis-muconate, and to regulate some of the same genes, such as catA and catB. Unlike CatM, BenM responded to benzoate. Benzoate together with cis,cis-muconate increased the BenM-dependent expression of the benABCDE operon synergistically. CatM was not required for this synergism, nor did CatM regulate the expression of a chromosomal benA::lacZ transcriptional fusion. BenM-mediated regulation differs significantly from that of the TOL plasmid-encoded conversion of benzoate to catechol in pseudomonads. The benM gene is immediately upstream of, and divergently transcribed from, benA, and a possible DNA binding site for BenM was identified between the two coding regions. Two mutations in the predicted operator/promoter region rendered ben gene expression either constitutive or inducible by cis,cis-muconate but not benzoate. Mutants lacking BenM, CatM, or both of these regulators degraded aromatic compounds at different rates, and the levels of intermediary metabolites that accumulated depended on the genetic background. These studies indicated that BenM is necessary for ben gene expression but not for expression of the cat genes, which can be regulated by CatM. In a catM-disrupted strain, BenM was able to induce higher levels of catA expression than catB expression.

Acinetobacter↗
PubMed1999

The genes rubA and rubB for alkane degradation in Acinetobacter sp. strain ADP1 are in an operon with estB, encoding an esterase, and oxyR.

Alkanes are oxidized in Acinetobacter sp. strain ADP1 by a three-component alkane monooxygenase, composed of alkane hydroxylase, rubredoxin, and rubredoxin reductase. rubA and rubB encode rubredoxin and a NAD(P)H-dependent rubredoxin reductase. We demonstrate here that single base pair substitutions in rubA or rubB lead to defects in alkane degradation, showing that both genes are essential for alkane utilization. Differences in the degradation capacity for hexadecane and dodecane in these mutants are discussed. Two genes, estB and oxyR, are located downstream of rubB, but are not necessary for alkane degradation. estB encodes a functional esterase. oxyR encodes a LysR-type transcriptional regulator, conferring resistance to hydrogen peroxide. rubA, rubB, estB, and oxyR constitute an operon, which is constitutively transcribed from a sigma70 promoter, and an estB-oxyR containing message is also transcribed from an internal promoter.

Acinetobacter↗
PubMed2000

The malonate decarboxylase operon of Acinetobacter calcoaceticus KCCM 40902 is regulated by malonate and the transcriptional repressor MdcY.

A regulatory gene-like open reading frame oriented oppositely to mdcL, coined mdcY, was found upstream from the structural genes of the mdcLMACDEGBH operon in Acinetobacter calcoaceticus KCCM 40902. To elucidate the function of this gene, mdcY was expressed in Escherichia coli, and the MdcY protein was purified to homogeneity. Its DNA binding activity and binding site were examined by gel retardation and footprinting assays in vitro and by site-directed mutagenesis of the binding sites in vivo. The regulator bound target DNA regardless of the presence of malonate, and the binding site was found centered at -65 relative to the mdcL transcriptional start site and contains a 12-bp palindromic structure (5'-ATTGTA/TACAAT-3'). Using a promoter fusion to the reporter gene luc, we found that the promoter P(mdcY) is negatively regulated by MdcY independent of malonate. However, the promoter P(mdcL) recovered its activity in the presence of malonate. When mdcY was introduced into A. calcoaceticus KCCM 40902 in which the gene is inactivated by an IS3 family element, malonate decarboxylase was significantly repressed in cultures growing in acetate, succinate, or Luria-Bertani medium. However, in cells growing in malonate, malonate decarboxylase was induced, indicating that MdcY is a transcriptional repressor and that malonate or a product resulting from malonate metabolism should be the intracellular inducer of the mdc operon.

Acinetobacter calcoaceticus↗