Showing papers on "Pseudomonas putida published in 1981"

Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta cleavage pathway

Abstract: The genetic organization of the Pseudomonas putida plasmid pWWO-161, which encodes enzymes for the degradation of toluene and related aromatic hydrocarbons, has been investigated by transposition mutagenesis and gene cloning. Catabolic genes were localized to two clusters, one for upper pathway (hydrocarbon leads to carboxylic acid) enzymes and the other for lower pathway (carboxylic acid leads to tricarboxylic acid cycle) enzymes, that are separated by a 14-kilobase DNA segment. The physical organization of the catabolic genes thus reflects their functional organization into two regulatory blocks. The pWWO-161 DNA fragments Sst I fragment C and fragment D were cloned in a broad host range vector to produce plasmid pKT530. This hybrid encodes toluate oxygenase and all meta cleavage pathway enzymes, and it enables P. putida mt-2 and Escherichia coli K-12 cells to grow on m-toluate as sole carbon source. The pKT530 plasmid also carries xylS (a gene whose product has been postulated to regulate expression of the lower pathway genes) and the control sequences of the pathway that interact with this product, because catechol 2,3-oxygenase synthesis is specifically induced by m-toluate in both P. putida and E. coli. Evidence is presented that suggests the promoter operator of the meta pathway gene functions less effectively with the RNA polymerase or xylS product of E. coli than with the enzyme or product of P. putida.

Properties of six pesticide degradation plasmids isolated from Alcaligenes paradoxus and Alcaligenes eutrophus.

Abstract: Biophysical and genetic properties of six independently isolated plasmids encoding the degradation of the herbicides 2,4-dichlorophenoxyacetic acid and 4-chloro-2-methylphenoxyacetic acid are described. Four of the plasmids, pJP3, pJP4, pJP5, and pJP7, had molecular masses of 51 megadaltons, belonged to the IncP1 incompatibility group, and transferred freely to strains of Escherichia coli, Rhodopseudomonas sphaeroides, Rhizobium sp., Agrobacterium tumefaciens, Pseudomonas putida, Pseudomonas fluorescens, and Acinetobacter calcoaceticus. In addition, these four plasmids conferred resistance to merbromin, phenylmercury acetate, and mercuric ions, had almost identical restriction endonuclease cleavage patterns, and encoded degradation of m-chlorobenzoate. The two other plasmids, pJP2 and pJP9, did not belong to the IncP1 incompatibility group, had molecular masses of 37 megadaltons, encoded the degradation of phenoxyacetic acid, and possessed identical restriction endonuclease cleavage patterns.

Inhibition of catechol 2,3-dioxygenase from Pseudomonas putida by 3-chlorocatechol.

Abstract: Partially purified preparations of catechol 2,3-dioxygenase from toluene-grown cells of Pseudomonas putida catalyzed the stoichiometric oxidation of 3-methylcatechol to 2-hydroxy-6-oxohepta-2,4-dienoate. Other substrates oxidized by the enzyme preparation were catechol, 4-methylcatechol, and 4-fluorocatechol. The apparent Michaelis constants for 3-methylcatechol and catechol were 10.6 and 22.0 muM, respectively. Substitution at the 4-position decreases the affinity and activity of the enzyme for the substrate. Catechol 2,3-dioxygenase preparations did not oxidize 3-chlorocatechol. In addition, incubation of the enzyme with 3-chlorocatechol led to inactivation of the enzyme. Kinetic analyses revealed that both 3-chlorocatechol and 4-chlorocatechol were noncompetitive or mixed-type inhibitors of the enzyme. 3-Chlorocatechol (Ki = 0.14 muM) was a more potent inhibitor than 4-chlorocatechol (Ki = 50 muM). The effect of the ion-chelating agents Tiron and o-phenanthrolene were compared with that of 3-chlorocatechol on the inactivation of the enzyme. Each inhibitor appeared to remove iron from the enzyme, since inactive enzyme preparations could be fully reactivated by treatment with ferrous iron and a reducing agent.

Isolation of alginate-producing mutants of Pseudomonas fluorescens, Pseudomonas putida and Pseudomonas mendocina.

Abstract: Spontaneous alginate-producing (muc) variants were isolated from strain of Pseudomonas fluorescens, P. putida and P. mendocina at a frequency of 1 in 108 by selcting for carbenicillin resistance. The infrared spectrum of the bacterial exopolysaccharide was typical of an acetylated alginate similar to that previously described in Azotobacter vinelandil and in mucoid variants of P. aeruginosa. Mucoid variants were not isolated from P. stuteri, P. pseudoalcaligenes, P. testosteroni, P. diminuta, P. acidovorans, P. cepacia or P. maltophilia.

Plasmid specifying total degradation of 3-chlorobenzoate by a modified ortho pathway.

Abstract: A plasmid, termed pAC25, specifying biodegradation of 3-chlorobenzoate in a strain of Pseudomonas putida has been characterized. During growth of the plasmid-harboring cells with 3-chlorobenzoate, there was an accumulation of 3-chlorocatechol and beta-chloromuconic acid as intermediates and release of more than 80% of the chlorine in the form of inorganic chloride. The plasmid had a mean molecular mass of 68 x 10(6) daltons and was transmissible to a number of Pseudomonas species such as P. aeruginosa, P. putida strain PpG1, and P. putida strain PRS1. Transfer of pAC25 to various catechol-negative mutants of P. putida strain PRS1 showed that the chromosomally coded pyrocatechase was not complemented by the plasmid-specified pyrocatechase, which appeared to be specific for the chlorinated catechols. In contrast to benzoate, which was metabolized by the ortho pathway through beta-ketoadipate as an intermediate, the plasmid specified ortho cleavage of the chlorocatechols through maleylacetate as an intermediate.

Catabolism of pseudocumene and 3-ethyltoluene by Pseudomonas putida (arvilla) mt-2: evidence for new functions of the TOL (pWWO) plasmid.

Abstract: Pseudocumene (1,2,4-trimethylbenzene) and 3-ethyltoluene were found to serve as growth substrates for Pseudomonas putida (arvilla) mt-2, in addition to toluene, m-xylene, and p-xylene as previously described. Similar observations were made with several additional P. putida strains also capable of growth with toluene and the xylenes. Additional substrates which supported the growth of these organisms included 3,4-dimethylbenzyl alcohol, 3,4-dimethylbenzoate, and 3-ethylbenzoate. P. putida mt-2 cells grown either with toluene or pseudocumene rapidly oxidized toluene, pseudocumene, and 3-ethyltoluene as well as 3,4-dimethylbenzoate, 3-ethylbenzoate, 3,4-dimethylcatechol, and 3-ethylcatechol. Cell extracts from similarly grown P. putida mt-2 cells catalyzed a meta fission of 3,4-dimethylcatechol and 3-ethylcatechol to compounds having the spectral properties of 2-hydroxy-5-methyl-6-oxo-2,4-heptadienoate and 2-hydroxy-6-ox-2,4-octadienoate, respectively. The further metabolism of these intermediates was shown to be independent of oxidized nicotinamide adenine dinucleotide (NAD+) and resulted in the formation of essentially equimolar amounts of pyruvate, indicating that each ring fission product was degraded via the hydrolytic branch of the meta fission pathway. Treatment of cells with N-methyl-N'-nitro-N-nitrosoguanidine led to the isolation of a mutant, which when grown with succinate in the presence of pseudocumene or 3-ethyltoluene accumulated 3,4-dimethylcatechol or 3-ethylcatechol. Cells unable to utilize toluene, m-xylene, and p-xylene, obtained by growth in benzoate, also lost the ability to utilize pseudocumene and 3-ethyltoluene. The ability to utilize these substrates could be reacquired by incubation with a leucine auxotroph otherwise able to grow on all of the aromatic substrates.

Isolation and Properties of 7β-(4-Carboxybutanamido)cephalosporanic Acid Acylase-producing Bacteria

Abstract: A search was undertaken for microorganisms that produce an enzyme capable of deacylating 7β-(4-carboxybutanamido)cephalosporanic acid (glutaryl 7-ACA, GL-7-ACA) to 7-aminocephalo-sporanic acid. Pseudomonas putida ATCC 950 was selected from our stock cultures, and Pseudomonas SY-77-1 was isolated from soil. Adding glutaric acid to the growth media enhanced the production of GL-7-ACA acylase by Pseudomonas SY-77-1. A β-lactamase-deficient mutant, Pseudomonas NS-2, was derived from Pseudomonas SY-77-1 treated with a mutagen. The cell suspension of Pseudomonas NS-2 specifically hydrolyzed cephalosporin compounds having aliphatic dicarboxylic acid in the acyl side chain.

Cyclic adenosine 3',5'-monophosphate levels in Pseudomonas putida and Pseudomonas aeruginosa during induction and carbon catabolite repression of histidase synthesis.

Abstract: Inducibility of histidase (histidine ammonia-lyase, EC 4.3.1.3) in Pseudomonas putida and Pseudomonas aeruginosa was observed to be strongly affected by succinate-provoked catabolite repression, but this did not occur as a consequence of reduced intracellular cyclic adenosine 3',5'-monophosphate levels, and repression could not be alleviated by exogenously added cyclic adenosine 3,'5'-monophosphate. Milder repression of histidase by lactate was also not reversed by the addition of cyclic adenosine 3',5'-monophosphate. These results, along with data showing intracellular cyclic adenosine 3',5'-monophosphate levels remained essentially constant during growth on such diverse carbon sources as histidine, acetamide, glucose, and succinate, indicated that catabolite repression of histidase synthesis by efficient carbon sources was not mediated through variations in internal cyclic adenosine 3,'5'-monophosphate.

Purification of a branched-chain keto acid dehydrogenase from Pseudomonas putida.

Abstract: We purified branched-chain keto acid dehydrogenase to a specific activity of 10 mumol/min per mg of protein from Pseudomonas putida grown on valine. The purified enzyme was active with 2-ketoisovalerate, 2-ketoisocaproate, and 2-keto-3-methylvalerate in a ratio of 1.0:0.8:0.7 but showed no activity with either pyruvate or 2-ketoglutarate. There were four polypeptides in the purified enzyme (molecular weights, 49,000, 46,000, 39,000, and 37,000). The purified enzyme was deficient in the specific lipoamide dehydrogenase produced during growth on valine (molecular weight, 49,000). Branched-chain keto acid dehydrogenase required L-valine, oxidized nicotinamide adenine dinucleotide, coenzyme A, thiamine pyrophosphate, and magnesium chloride. A partially purified preparation catalyzed the oxidation of 2-keto-[1-14C]isovalerate to [14C]carbon dioxide, isobutyryl-coenzyme A, and reduced nicotinamide adenine dinucleotide in equimolar amounts. Both the Km and the Vmax for 2-ketoisovalerate were affected by the addition of L-valine to the assay mixture. However, only the Vmax values for oxidized nicotinamide adenine dinucleotide and coenzyme A were affected when L-valine was present. This suggested that valine acted by affecting the binding of branched-chain keto acids to subunit E1 of the complex.

Isolation and characterization of spontaneously occurring TOL plasmid mutants of Pseudomonas putida HS1.

Abstract: A strain of Pseudomonas (P. putida HS1) was found to resemble P. putida (arvilla) mt-2 in its ability to degrade toluene, m- and p-xylene, 1,2,4-trimethylbenzene (pseudocumene), and 3-ethyltoluene via oxidation of a methyl substituent and reactions of the meta-fission pathway. The ability to degrade these substrates by P. putida HS1 (PpC1) was shown to be encoded by a TOL (pDK1) plasmid as evidenced by: (i) spontaneous loss of the TOL-related phenotype after growth with benzoate, (ii) transfer of the TOL character from the wild type into cured recipients by conjugation, and (iii) isolation of a plasmid of identical molecular weight (120 X 10(6)) from both the wild type and an exconjugant obtained by mating wild type with a putative cured recipient. In addition to the isolation of apparent cured strains having lost the entire TOL-related phenotype, two additional mutant classes were observed after growth on benzoate. One class, represented by PpCT1, was unable to utilize the alkyl-substituted aromatic compounds but retained the ability to grow with toluene and benzyl alcohol. Analysis of PpCT1 revealed that it was unable to synthesize the TOL-encoded toluate oxidase and enzymes of the meta pathway but retained the ability to elaborate activities for toluene hydroxylase, benzyl alcohol, and benzaldehyde dehydrogenase, thereby mediating initial oxidation of toluene to benzoate, which was then further metabolized via enzymes of the chromosomally encoded ortho-fission pathway. A second class of mutants had lost the ability to utilize the hydrocarbons but could still grow with m-toluate but not p-toluate, 3,4-dimethylbenzoate, or 3-ethylbenzoate, intermediates in the oxidation of the corresponding hydrocarbons. Our such mutant, PpCM1, could no longer synthesize enzymes required for initial oxidation of the hydrocarbons, but was able to produce the toluate oxidase and enzymes of the meta pathway, thereby facilitating degradation of m-toluate. Neither PpCT1, PpCM1, nor a putative cured strain, PpCC1, reverted at detectable frequencies (less than 10(-9). Analysis of each strain for plasmid deoxyribonucleic acid revealed the presence of a single plasmid in each strain with the following molecular weights: PpCM1, 100 X 10(6) (pDKM1); PpCT1, 80 X 10(6) (pDKT1); PpCC1 20 X 10(6) (pDKC1). The results suggest that the TOL (pDK1) plasmid has undergone deletions giving rise to smaller replicons which either encode for only a fraction of the wild-type catabolic functions (pDKM1, pDKT1) or have lost all catabolic activities (pDKC1).

8 alpha-(O-Tyrosyl)flavin adenine dinucleotide, the prosthetic group of bacterial p-cresol methylhydroxylase.

Abstract: 8 alpha-(O-Tyrosyl)riboflavin has been synthesized by condensation of the copper complex of L-tyrosine with 8 alpha-bromotetraacetylriboflavin. The structure of this synthetic product was proven by absorption and 1H NMR spectroscopy and by chemical degradation, which yielded 1 mol of tyrosine per mol of flavin. The synthetic compound comigrated wtih the (aminoacyl)riboflavin isolated from the p-cresol methylhydroxylase of Pseudomonas putida, and both showed identical absorption and fluorescence spectral properties. 8 alpha-(O-Tyrosyl)riboflavin as well as the flavin-containing decapeptide from p-cresol methylhydroxylase undergoes reductive cleavage to form riboflavin and FAD, respectively, on anaerobic treatment with dithionite. In contrast, the native enzyme, on reduction with dithionite, yields a reduced flavin via a red (anionic) flavosemiquinone intermediate, which remains covalently bound to the protein even under denaturing conditions. 8 alpha-(O-Tyrosyl)riboflavin bound to apoflavodoxin is also not cleaved on reduction with dithionite, but, instead, a blue (neutral) semiquinone of tyrosylriboflavin is generated, which is resistant to further reduction with dithionite. Three p-cresol methylhydroxylases, isolated from different strains of Pseudomonas putida, differing in molecular weight and Km values for substrates, contain the same peptide at the flavin site. These data provide definitive proof for the existence of 8 alpha-(O-tyrosyl)riboflavin in nature.

Isolation of a specific lipoamide dehydrogenase for a branched-chain keto acid dehydrogenase from Pseudomonas putida.

Abstract: We purified lipoamide dehydrogenase from cells of Pseudomonas putida PpG2 grown on glucose (LPD-glu) and lipoamide dehydrogenase from cells grown on valine (LPD-val), which contained branched-chain keto acid dehydrogenase. LPD-glu had a molecular weight of 56,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and LPD-val had a molecular weight of 49,000. The pH optimum for LPD-glu for reduced nicotinamide adenine dinucleotide oxidation was 7.4, compared with pH 6.5 for LPD-val. When oxidized nicotinamide adenine dinucleotide was included in the assay mixture, the pH optima were 7.1 and 5.7, respectively. There was also a difference in pH optima between the two enzymes for oxidized nicotinamide adenine dinucleotide reduction, but the Michaelis constants and maximum velocities were similar. A purified preparation of branched-chain keto acid dehydrogenase, which was deficient in lipoamide dehydrogenase, was stimulated 10-fold by LPD-val but not by LPD-glu, which suggested that the branched-chain keto acid dehydrogenase of P. putida has a specific requirement for LPD-val. In contrast, a partially purified preparation of 2-ketoglutarate dehydrogenase that was deficient in lipoamide dehydrogenase was stimulated by LPD-glu but not by LPD-val, indicating that this complex has a specific requirement of LPD-glu. Images

Plasmid- and chromosome-mediated dissimilation of naphthalene and salicylate in Pseudomonas putida PMD-1.

Abstract: Pseudomonas putida PMD-1 dissimilates naphthalene (Nah), salicylate (Sal), and benzoate (Ben) via catechol which is metabolized through the meta (or alpha-keto acid) pathway. The ability to utilize salicylate but not naphthalene was transferred from P. putida PMD-1 to several Pseudomonas species. Agarose gel electrophoresis of deoxyribonucleic acid (DNA) from PMD-1 and Sal+ exconjugants indicated that a plasmid (pMWD-1) of 110 megadaltons is correlated with the Sal+ phenotype; restriction enzyme analysis of DNA from Sal+ exconjugants indicated that plasmid pMWD-1 was transmitted intact. Enzyme analysis of Sal+ exconjugants demonstrated that the enzymes required to oxidize naphthalene to salicylate are absent, but salicylate hydroxylase and enzymes of the meta pathway are present. Thus, naphthalene conversion to salicylate requires chromosomal genes, whereas salicylate degradation is plasmid encoded. Comparison of restriction digests of plasmid pMWD-1 indicated that it differs considerably from the naphthalene and salicylate degradative plasmids previously described in P. putida.

Bacterial degradation of 3,4,5-trimethoxycinnamic acid with production of methanol.

Abstract: When grown on 3,4,5-trimethoxycinnamic acid, a strain of Pseudomonas putida oxidized this compound and also 3,4,5-trimethoxybenzoic, 3,5-dimethoxy-4-hydroxybenzoic (syringic), and 3,4-dihydroxy-5-methoxybenzoic (3-O-methylgallic) acids, but 3,5-dimethoxy-4-hydroxycinnamic and other acids bearing structural resemblances to the growth substrate were oxidized only slowly. These results indicate that two carbon atoms of the side chain of 3,4,5-trimethoxycinnamate were released before oxidative demethylation occurred to give the ring-fission substrate, 3-O-methylgallate. Oxidation of 3,4,5-trimethoxycinnamate by intact cells gave equimolar amounts of methanol, which was derived from the methoxyl group of 3-O-methylgallate. The tricarboxylic acids, 4-carboxy-2-keto-3-hexenedioic and 4-carboxy-4-hydroxy-2-ketoadipic acids, were shown to be formed by the action of a cell extract upon 3-O-methylgallate; therefore, methanol was released either during or immediately after fission of the benzene nucleus. Cell extracts, prepared from several independent soil isolates after growth on 3,4,5-trimethoxy derivatives of benzoic, cinnamic, and beta-phenylpropionic acids, rapidly oxidized 3-O-methylgallate without added cofactors. It is concluded that the reactions investigated serve generally as a source of methanol in nature.

Mössbauer spectroscopic studies of the terminal dioxygenase protein of benzene dioxygenase from Pseudomonas putida.

Abstract: Mossbauer spectra obtained from the terminal dioxygenase protein of the benzene dioxygenase system from Pseudomonas putida show that it contains [2Fe--2S] centres similar to those of the two-iron plant-type ferredoxins. In the oxidized form the two iron atoms within the centre are high-spin ferric but with considerable inequivalence. In the reduced form the centre contains one extra electron, and this is localized on one of the iron atoms, which becomes high-spin ferrous.

Regulation of the utilization of glucose and aromatic substrates in four strains of Pseudomonas putida

Abstract: During the oxidation of various mixtures of glucose and aromatic substrates by four strains of Pseudomonas putida, diauxic growth was not observed. Strain A3.12 grew faster on benzoate than on glucose, whereas three other strains showed faster growth on glucose than on the aromatic test substrates. Growth rates on mixtures of glucose and aromatics were intermediate between those on the single substrates. The presence of glucose in media containing aromatic substrates accelerated in the bacteria the appearance of the ability to oxidize aromatic substrates. During growth of the organisms on binary mixtures of aromatics, simultaneous utilization of these compounds occurred, the utilization ratio depending on the quality of the compounds as carbon and energy sources. Addition of glucose to dual aromatic substrate media greatly increased the utilization ratio in favour of the “better” aromatic substrate. With decreasing concentration of glucose in relation to that of aromatic substrates, the rate of carbon assimilation from glucose increased. Enzymological and radiochemical studies demonstrated that even in the presence of an excess of aromatic substrates, glucose was exclusively catabolized via the 2-keto-3-deoxy-6-phosphogluconate pathway. In contrast, the rate of carbon assimilation from 14C-ring-labelled benzoate and anisate was unaffected by the presence of an excess of glucose.

Isolation from legume tissues of an agglutinin of saprophytic pseudomonads

Abstract: Water extracts from leaves, roots and stems of green bean and leaves of mung bean, soybean, and lima bean caused agglutination of cells of a saprophytic bacterium, Pseudomonas putida. The agglutinin from each source was purified in an identical manner using procedures of ion exchange, gel, and affinity chromatography. Purified preparations contained both carbohydrate and protein, were of high molecular weight (at least 106), required Mg2+ for activity and were heat stable. Less than 1 μg of hexose or protein of the purified preparations was required to completely agglutinate about 109P. putida cells. Arabinose, galactose and galacturonic acid were the predominant glycosyl residues detected in the purified agglutinin preparations. Agglutination tests with the crude and purified preparations from each legume showed strong and consistent agglutination with only pseudomonad saprophytes, P. putida, P. fluorescens, and P. aeruginosa. No agglutination of other plant pathogenic bacteria or nonpseudomonad saprophy...

Glutamine synthetase of pseudomonads: some biochemical and physicochemical properties.

Abstract: The glutamine synthetases from several Pseudomonas species were purified to homogeneity, and their properties were compared with those reported for the enzymes from Escherichia coli and other gram-negative bacteria. The glutamine synthetase from Pseudomonas fluorescens was unique because it was nearly precipitated quantitatively as a homogeneous protein during dialysis of partially purified preparations against buffer containing 10 mM imidazole (pH 7.0) and 10 mM MnCl2. The glutamine synthetases from Pseudomonas putida and Pseudomonas aeruginosa were purified by affinity chromatography on Affi-blue gel. Dodecamerous forms of the E. coli and P. fluorescens glutamine synthetases had identical mobilities during polyacrylamide gel electrophoresis. Their dissociated subunits, however, migrated differently and were readily separated by electrophoresis on polyacrylamide gels containing 0.1% sodium dodecyl sulfate. This difference in subunit mobilities is not related to the state of adenylylation. Regulation of the Pseudomonas glutamine synthetase activity is mediated by an adenylylation-deadenylylation cyclic cascade system. A sensitive procedure was developed for measuring the average number of adenylylated subunits per enzyme molecule for the glutamine synthetase from P. fluorescens. This method takes advantage of the large differences in transferase activity of the adenylylated and unadenylylated subunits at pH 6.0 and of the fact that the activities of both kinds of subunits are the same at pH 8.45.

Regulation of phenol degradation in Pseudomonas putida.

Abstract: In order to characterize the ability of Pseudomonas putida (TREVISAN 1889) MIGULA 1895 strain H to degrade various mono- and diphenolic aromatic compounds, respiratory activities towards phenol, catechol, and the cresol isomers were determined. The following rates of oxygen uptake (QO2) were obtained with resting phenol-grown cells: phenol -- 229, o-cresol -- 231, m-cresol -- 43, p-cresol -- 200, catechol -- 262. All these compounds were oxidized by a two-phase-kinetics, the first phase is characterized by a higher oxidation rate than the second. The oxidation of phenol as well as of p-cresol was found to be substrate-inhibited at concentrations above 0.25 mM. A Ki-value of 100 mg/l was calculated for phenol oxidation. The phenol-degrading enzyme system is induced, probably coordinately, by phenol and the cresol isomers. In strain H the degradation of phenol is carried out simultaneously with the assimilation of natural carbohydrates like glucose and sodium pyruvate. Aniline as well as sodium benzoate, though not metabolized by strain H, cause a concentration-dependent inhibition of phenol degradation in resting phenol-grown cells of that strain. The mechanism of this inhibition is discussed.

Production of muconic acid

Abstract: This invention provides a process for microbiological oxidation of toluene to muconic acid. The toluene oxidation is achieved with novel strains of microorganisms (e.g., Pseudomonas putida Biotype A) which are capable of converting toluene to muconic acid quantitatively by the ortho (β-ketoadipate) pathway. Munonate lactonizing enzyme is not induced in the microorganism, thereby permitting the muconic acid to be produced and accumulated in a quantity greater than one gram of muconic acid per liter of conversion medium.

In Vitro Antimicrobial Activity of Ceftizoxime Against Glucose-Nonfermentative Gram-Negative Rods

Abstract: Ceftizoxime, a new cephalosporin, was active against Pseudomonas cepacia, Flavobacterium meningosepticum, Alcaligenes faecalis, and Acinetobacter calcoaceticus and was more potent against Pseudomonas aeruginosa and Pseudomonas putida than was carbenicillin.

Control of microorganisms in the rhizosphere of wheat by inoculation of seeds with Pseudomonas putida and by foliar application of urea

Abstract: After inoculation of wheat seeds with various bacterial strains germination of plants was usually inhibited at first but growth was stimulated later. After inoculation withPseudomonas putida K 11 producing physiologically active compounds the total number of bacteria increased together with the bacteria: fungi ratio in the rhizosphere. These characteristic were further increased after foliar application of urea due to increased root exudation. Dry mass of upper wheat parts was about 15 — 80 % higher in green-house experiments, in which the plants were treated in the two above ways. More reliable results, were usually obtained by bacterization ofP. putida and foliar application of urea as compared with the situation when the seeds were inoculated without the foliar application or, on the contrary, after foliar application without inoculation of the seeds. Only when urea was applied early and in a soil contaminated with the fungusGaeumannomyces graminis var.tritici (causing “take-all” of the wheat) no favourable results could be detected. In these cases the foliar application without inoculation of the seeds was more successful. Symptoms of the disease of wheat roots caused byG. graminis were less frequently observed after the inoculation of the seeds with the strainP. putida K 11 and after the foliar application of urea.

Control of meta-cleavage degradation of 4-hydroxyphenylacetate in Pseudomonas putida.

Abstract: Synthesis of enzymes of the 4-hydroxyphenylacetate meta-cleavage pathway was studied in Pseudomonas putida wild-type strain P23X1 (NCIB 9865) and mutant strains which had either structural or regulatory gene mutations. Induction studies with mutant strains each defective in an enzyme of the pathway showed that 4-hydroxyphenylacetate induced the hydroxylase and that 3,4-dihydroxyphenylacetate induced the 2,3-oxygenase, aldehyde dehydrogenase, isomerase, decarboxylase, and hydratase. This showed that the hydroxylase structural gene does not exist in an operon that contains any other structural gene of this meta pathway. Studies of mutant strains that synthesized constitutively the 2,3-oxygenase and subsequent enzymes suggested that the regulation of synthesis of these enzymes was coincident, and, in such strains, the hydroxylase was inducible only. Observations made with a putative polarity mutant that lacked 2,3-oxygenase activity suggested that the structural genes encoding this enzyme and subsequent enzymes of the pathway exist in the same operon. Studies of a regulatory mutant strain that was defective in the induction of the 2,3-oxygenase and subsequent enzymes suggest that the 2,3-oxygenase operon is under positive control.

Process for treating wastewater containing phenolics and microorganism capable of degrading phenolics

Abstract: Mutant microorganism, Pseudomonas putida CB-173 degrading phenolics, and at a temperature as low as, e.g., about 1° to 4° C., at a faster rate than known Pseudomonas putida type strains, and process for treating wastewater containing phenolics using the mutant microorganism strain Pseudomonas putida CB-173.