| extraction of a steroid transport system from pseudomonas testosteroni membranes and incorporation into synthetic liposomes. | a steroid binding protein has been extracted from pseudomonas testosteroni membranes with an organic solvent system. this protection binds some c19 and c21 steroids but not c18 steroids. when this protein is incorporated into synthetic lipid vesicles constructed from p. testosteroni phospholipids, the vesicles perform concentrative uptake of testosterone in the presence of the ionophore valinomycin. this steroid binding protein is thus believed to be the steroid permease of this organism. | 1985 | 4068713 |
| use of a solid-phase photoaffinity reagent to label a steroid binding site: application to the delta 5-3-ketosteroid isomerase of pseudomonas testosteroni. | in order to extend our analysis of the reactions that occur during the active site directed photoinactivation of delta 5-3-ketosteroid isomerase sensitized by unsaturated steroid ketone photoaffinity reagents, the site of covalent attachment has been identified. a solid-phase photoaffinity reagent, delta 6-testosterone-agarose, has been employed for this purpose; this type of reagent, in contrast to solution-phase reagents, facilitated the recovery of a peptide fragment of the isomerase bearing ... | 1985 | 4092021 |
| [determination of the characteristics of an inductor necessary for its interaction with a repressor in pseudomonas testosteroni]. | | 1972 | 4119671 |
| on the 3 alpha-hydroxysteroid dehydrogenase from pseudomonas testosteroni. purification and properties. | | 1974 | 4137094 |
| degradation of protocatechuate in pseudomonas testosteroni by a pathway involving oxidation of the product of meta-fission. | in addition to catalyzing the hydrolysis of 4-carboxy-2-hydroxymuconic semialdehyde, formed by meta-fission of protocatechuate, pseudomonas testosteroni also possesses a nicotinamide adenine dinucleotide(phosphate)-linked dehydrogenase for this compound and can degrade protocatechuate to pyruvate and oxaloacetate. | 1973 | 4143957 |
| the degradation of l-histidine, imidazolyl-l-lactate and imidazolylpropionate by pseudomonas testosteroni. | 1. imidazol-5-ylpropionate and imidazol-5-yl-lactate are degraded by pseudomonas testosteroni via inducible pathways. 2. growth on either compound as the sole source of carbon results in the induction of the enzymes for histidine catabolism. 3. the pathway of histidine degradation in this organism, a non-fluorescent pseudomonad, is shown to be the same as that operating in pseudomonas fluorescens and pseudomonas putida. it consists of the successive formation of urocanate, imidazol-4-on-5-ylprop ... | 1973 | 4146796 |
| the control of the enzymes degrading histidine and related imidazolyl derivates in pseudomonas testosteroni. | 1. the induction of the enzymes for the degradation of l-histidine, imidazolylpropionate and imidazolyl-l-lactate in pseudomonas testosteroni was investigated. 2. the activities of histidine ammonia-lyase, histidine-2-oxoglutarate aminotransferase and urocanase are consistent with these enzymes being subject to co-ordinate control under most growth conditions. however, a further regulatory mechanism may be superimposed for histidase alone under conditions where degradation of histidine must take ... | 1973 | 4146797 |
| 3-hydroxybenzoate 4-hydroxylase from pseudomonas testosteroni. | | 1973 | 4148586 |
| reversibility of steroid delta-isomerase. 3. the soluble enzyme of pseudomonas testosteroni. | | 1966 | 4287910 |
| [contribution to the study of the active site of 3 (or 17)-beta-hydroxysteroid: nad oxidoreductase of pseudomonas testosteroni]. | | 1967 | 4293245 |
| [on the inhibition by excess substrate of beta-hydroxysteroid dehydrogenase from pseudomonas testosteroni]. | | 1968 | 4301944 |
| absolute configuration of a metabolite in the m-fission pathway of protocatechuate. | an aldolase, which is induced in pseudomonas testosteroni during growth with p-hydroxybenzoate, preferentially attacks the r form of 4-hydroxy-4-methyl-2-oxoglutarate, a metabolite of protocatechuate catabolism. | 1973 | 4347922 |
| steroid-receptor in pseudomonas testosteroni released by osmotic shock. | | 1973 | 4363616 |
| induction of steroid-binding activity in pseudomonas testosteroni. | | 1973 | 4363617 |
| affinity chromatography of 3-oxosteroid delta4--delta5-isomerase of pseudomonas testosteroni. | | 1974 | 4367971 |
| periplasmic steroid-binding proteins and steroid transforming enzymes of pseudomonas testosteroni. | | 1974 | 4376195 |
| [enzyme induction with steroids in pseudomonas testosteroni]. | | 1965 | 4379247 |
| [inhibition, with alkaloids, of the steroid-dependent enzyme induction in pseudomonas testosteroni]. | | 1967 | 4384350 |
| enzymatic determination of bile acids. the presence of a new enzyme, a 12alpha-hydroxysteroid: nad-oxidoreductase in extracts from pseudomonas testosteroni. | | 1974 | 4422066 |
| amino acid sequence of a peptide containing the active cysteine residue of histidine ammonia-lyase. | 1. oxidized (polymerized) histidine ammonia-lyase from pseudomonas testosteroni was activated with dithiothreitol and the reduced disulphide-linked cysteine residues of the native enzyme were carboxymethylated with iodo[(14)c]acetate. 2. the activity of the carboxymethylated enzyme was similar to that of the polymerized form and approx. 15% of that of the fully reduced form. 3. a tryptic digest of the [(14)c]carboxymethylated enzyme contained only one radioactive peptide. 4. the amino acid seque ... | 1974 | 4422650 |
| proceedings: synthesis of rna and dna in isolated chromosome of pseudomonas testosteroni. | | 1974 | 4461578 |
| differentiation between pseudomonas testosteroni and p. acidovorans by gas chromatography. | | 1972 | 4561033 |
| [studies on rna polymerase dna complex from pseudomonas testosteroni]. | | 1972 | 4649824 |
| the regulation of the -ketoadipate pathway in pseudomonas acidovorans and pseudomonas testosteroni. | | 1972 | 4657135 |
| micellar aggregation of 5 -3-keto steroids lacking a polar c-17 group and its relation to the activity and specificity of the 5 - 4 -3-ketosteroid isomerase of pseudomonas testosteroni. | | 1973 | 4683484 |
| molecular weight determination and structural studies of pseudomonas testosteroni delta 5 leads to 4-3-oxosteroid isomerase (ec 5.3.3.1). | | 1973 | 4753764 |
| [relation between enzyme induction and testosterone metabolism in pseudomonas testosteroni]. | | 1973 | 4803170 |
| influence of an 18-hydroxyl group on the interaction of oestrogens and hydroxysteroid oxidoreductases. | 1. partially purified 17beta-hydroxy steroid-nad(+) oxidoreductases, prepared from pseudomonas testosteroni (ec 1.1.1.51), human term placenta (ec 1.1.1.62) and the cytoplasmic fraction of rat liver (ec 1.1.1.-) were tested for their ability to catalyse the oxidoreduction of 18-hydroxyoestradiol-17beta and 18-hydroxyoestrone. the products of incubation were identified by chromatographic procedures and by mass spectrometry. 2. the pseudomonas enzyme catalysed both the oxidation of 18-hydroxyoestr ... | 1974 | 4824210 |
| [inhibition of a dna-dependent rna biosynthesis by a macromolecular fraction of non-induced cells of pseudomonas testosteroni]. | | 1971 | 4939704 |
| inhibition of the 3-ketosteroid 5 - 4 -isomerase of pseudomonas testosteroni by some bromo-3-ketosteroid derivatives. | | 1972 | 5017704 |
| the amino acid sequence of 5 -3-ketosteroid isomerase of pseudomonas testosteroni. | | 1971 | 5135313 |
| enzymatic oxidation of steroids by cell-free extracts of pseudomonas testosteroni: isolation of cleavage products of ring a. | | 1965 | 5217462 |
| inhibition by 2'-deoxyadenosine of enzyme induction in pseudomonas testosteroni. | | 1965 | 5218718 |
| [relationship between the structure of a steroid and the inducing effect on 3 alpha-hydroxysteroid: nad oxidoreductase of pseudomonas testosteroni]. | | 1969 | 5345977 |
| bacterial metabolism of arylsulfonates. i. benzene sulfonate as growth substrate for pseudomonas testosteroni h-8. | pseudomonas testosteroni h-8 utilizes as sole carbon source benzene sulfonate (bs), p-toluene sulfonate (pts), and ethylbenzene sulfonate (ebs) but not higher homologs. growth on bs was rapid (generation time, 3 hr) and efficient (y = 57), and resulted in accumulation of sulfate. as the culture is acid-sensitive, the medium must be heavily buffered to permit extensive growth. the bs oxidase system is inducible. cells grown on bs, but not glutamate, oxidized bs, pts, or ebs without lag (qo(2) = 5 ... | 1971 | 5553286 |
| studies with the 3-alpha-hydroxysteroid dehydrogenase from pseudomonas testosteroni-enzyme-substrate complementarity as the basis of selectivity and steric specificity. | | 1967 | 5597320 |
| the metabolism of aromatic acids by pseudomonas testosteroni and p. acidovorans. | | 1967 | 5602468 |
| the metabolism of protocatechuate by pseudomonas testosteroni. | 1. protocatechuate 4,5-oxygenase, purified 21-fold from extracts of pseudomonas testosteroni, was examined in the ultracentrifuge and assigned a mol.wt. of about 140000. 2. when diluted, the enzyme rapidly lost activity during catalysis. inactivation was partially prevented by l-cysteine. 3. with a saturating concentration of protocatechuate (1.36mm), k(m) for oxygen was 0.303mm. this value is greater than the concentration of oxygen in water saturated with air at 20 degrees . 4. cell extracts c ... | 1968 | 5683506 |
| further studies of steoidal inhibitors of delpha5, 3beta-hydroxysteroid dehydrogenase and delta5-delta4, 3-ketosteroid isomerase in pseudomonas testosteroni and in bovine adrenals. | | 1968 | 5697036 |
| preparation of crystalline delta-5-3-ketosteroid isomerase from pseudomonas testosteroni. | | 1965 | 5849828 |
| inhibition of 3-beta-hydroxysteroid dehydrogenase from pseudomonas testosteroni by various estrogenic and progestinic steroids. | | 1967 | 6023578 |
| inhibition of the steroid-induced synthesis of delta-5-3-ketosteroid isomerase in pseudomonas testosteroni by a new purine deoxyribonucleoside analog: 6-chloro-8-aza-9-cyclopentylpurine. | | 1967 | 6033637 |
| [fatty acids of the cell wall and the membrane of pseudomonas testosteroni]. | | 1967 | 6053273 |
| protection of substrate against enzymatic action by binding to proteins. dependence upon enzyme and binding protein. | in fetal or adult rats estradiol is carried in the plasma by alpha-fetoprotein or albumin. the protection of the carriers toward enzymatic oxidation by 17 beta-hydroxysteroid dehydrogenase from rat liver has been studied. concentrations of carrier protein and estradiol were adjusted to give free estradiol concentrations varying from km/10 to km/100 and the ratio of the catalytic velocity to that observed for the same concentration of free estradiol in the absence of carrier protein were recorded ... | 1980 | 6156164 |
| norethisterone and ethinylestradiol do not inhibit delta 5-3 beta-hydroxysteroid dehydrogenase in rat leydig cells. | the effect of norethisterone, norethisterone acetate, ethinylestradiol and of a cyanoketone on the activity of hsd have been studied. rat leydig cells and extracts of pseudomonas testosteroni were used as enzyme sources. conversion of [3h]dhea to a and [3h]pregnenolone to progesterone were used for the assay of enzyme activity. km values for each substrate for leydig cells and for bacterial enzyme were: 1.5 x 10(-5) mol l-1 and 6.0 x 10(-5) mol l-1 dhea; 1.3 x 10(-5) mol and 7.7 x 10(-5) mol l-1 ... | 1984 | 6237898 |
| influence of the position of the double bond in steroid substrates on the efficiency of the proton-transfer reaction by pseudomonas testosteroni 3-oxo-steroid delta 4-delta 5-isomerase. | studies of the proton-transfer reaction by pseudomonas testosteroni 3-oxo steroid delta(4)-delta(5)-isomerase with delta(5(6))- and delta(5(10))-steroid substrates demonstrate the importance of the position of the double bond for the efficiency of the isomerization process. thus 3-oxo-delta(5(6))-substrates have markedly high k(cat.) values, whereas those of 3-oxo-delta(5(10))-substrates are very low and their apparent k(m) values approach equilibrium dissociation constants. the first step in th ... | 1980 | 6248031 |
| proton nuclear magnetic resonance studies of pseudomonas testosteroni 3-oxo-delta 5-steroid isomerase and its interaction with 17 beta-estradiol. | | 1982 | 6295444 |
| the effect of carbonyl cyanide m-chlorophenylhydrazone on steroid transport in membrane vesicles of pseudomonas testosteroni. | the uncoupler carbonyl cyanide chlorophenylhydrazone (cccp) was an effective inhibitor of steroid transport in membrane vesicles of pseudomonas testosteroni between 10 microm and 1 microm cccp. at these concentrations the inhibition of steroid transport was not due to an inhibition of the 3 beta and 17 beta-hydroxysteroid dehydrogenase enzyme. cccp also affected testosterone-dependent oxygen consumption at concentrations up to 100 microm and inhibited respiration at 0.5 and 1 microm. the effect ... | 1983 | 6310264 |
| epr and mössbauer studies of protocatechuate 4,5-dioxygenase. characterization of a new fe2+ environment. | protocatechuate 4,5-dioxygenase from pseudomonas testosteroni has been purified to homogeneity and crystallized. the iron containing, extradiol dioxygenase is shown to be composed of two subunit types (alpha, mr = 17,700 and beta, mr = 33,800) in a 1:1 ratio; such a composition has not been observed for other extradiol dioxygenases. the 4.2 k mössbauer spectrum of native protocatechuate 4,5-dioxygenase prepared from cells grown in 57fe-enriched media consists of a doublet with quadrupole splitti ... | 1983 | 6317682 |
| [transformation of escherichia coli and different species of pseudomonas by a plasmid dna isolated from pseudomonas testosteroni]. | pseudomonas testosteroni uses testosterone as sole source of carbon. we were able to isolate an extrachromosomal dna from a strain of pseudomonas testosteroni and to obtain pseudomonas putida and aeruginosa and escherichia coli transformants catabolizing testosterone. | 1983 | 6416627 |
| [isolation and study of a bacteriophage of pseudomonas testosteroni]. | a virulent phage specific for pseudomonas testosteroni is described. this phage have a regular icosahedral head (52 nm between opposite angles) and a contractile tail (165 x 8 nm) but no fibers on. the buoyant density is 1,51 +/- 0,01 g/ml. the nucleic acid is an desoxyribonucleic acid with a density of 1,696 +/- 0,03 g/ml and a gc% between 33,7 and 39,7. | 1980 | 6451268 |
| evaluation of the rapid nft system for identification of gram-negative, nonfermenting rods. | this study evaluated the ability of the rapid nft system (api system sa, montalieu-vercieu, france) to accurately identify 262 clinically isolated, gram-negative, nonfermentative rods without additional tests. identifications were classified as correct; low discrimination, with a spectrum of two or more possibilities (additional tests necessary for accurate identification); and incorrect. correct identification rates were analyzed in two categories: (i) correct to species or biotype for all orga ... | 1984 | 6490857 |
| differences in inhibition by various steroids of rat testis and pseudomonas testosteroni delta 5-3 beta-hydroxysteroid dehydrogenase. | the influence of various estrogens, progestogens and of cyanoketone (2 alpha-cyano-4,4,17 alpha-trimethylandrost-5-en-17 beta-ol-3-one) on the enzyme activity of delta 5-3 beta-hydroxysteroid dehydrogenase (hsd) (ec 1.1.1.145) was studied. extracts of pseudomonas testosteroni, rat testis total homogenate and a microsomal preparation were used as enzyme sources. spectrophotometric determinations and the conversion of 3h-labelled dehydroepiandrosterone to androstenedione were used to assay for enz ... | 1983 | 6603965 |
| purification and characterization of a membrane-associated testosterone-binding protein from pseudomonas testosteroni. | a steroid-binding protein, identified in the supernatant generated when membrane vesicles of pseudomonas testosteroni are produced and harvested by centrifugation, has been purified 49-fold to homogeneity. it has a molecular weight of 30 000-35 000 and it specifically binds the c19 steroids dihydrotestosterone, testosterone, and androstenedione. it is a basic protein with an isoelectric point at ph 7.3. binding of testosterone exhibited normal saturation kinetics with an affinity constant, kd, o ... | 1983 | 6683990 |
| enzymatic function in crystals of delta 5-3-ketosteroid isomerase. catalytic activity and binding of competitive inhibitors. | crystals of the steroid-metabolizing enzyme, delta 5-3-ketosteroid isomerase (ec 5.3.3.1) from pseudomonas testosteroni, exhibit many enzymatic properties. each enzyme subunit in the lattice binds a competitive inhibitor, progesterone, with the same stoichiometry (1:1) and affinity (kd = 6 x 10(-6) m) as the enzyme in solution. another competitive inhibitor, 19-nortestosterone, competes with progesterone for the same binding sites in the crystal. the enzyme crystals catalyze the conversion of de ... | 1984 | 6746640 |
| antibodies to pseudomonas testosteroni 3-oxosteroid delta 4-delta 5-isomerase. | | 1980 | 6774719 |
| the effect of 2,4-dinitrophenol on steroid transport in membrane vesicles of pseudomonas testosteroni. | steroid transport in pseudomonas testosteroni membrane vesicles was significantly inhibited by the uncoupled 2,4-dinitrophenol (dnp). inhibition of steroid transport was not due to inhibition of the 3 beta- and 17 beta-hydroxysteroid dehydrogenase by concentrations of up to 1 mm dnp. however, inhibition of this membrane-bound enzyme was measured at 10 mm dnp. the solubilized 3 beta- and 17 beta-hydroxysteroid dehydrogenase was more sensitive, being inhibited at both 1 and 10 mm dnp indicating a ... | 1983 | 6827840 |
| photoaffinity modification of delta 5-3-ketosteroid isomerase by light-activatable steroid ketones covalently coupled to agarose beads. | in order to identify the minor site(s) of photoattachment of unsaturated steroid ketones to delta 5-3-ketosteroid isomerase from pseudomonas testosteroni, we have developed a solid-state photoaffinity labeling technique. two solid-state reagents, o-carboxymethylagarose-ethylenediamine-succinyl-17 beta-o-19-nortestosterone and o-carboxymethylagarose-ethylenediamine-succinyl-17 beta-o-4,6-androstadien-3-one, have been synthesized. under anaerobic conditions, isomerase bound to these resins is phot ... | 1983 | 6860646 |
| immunological relatedness of histidine ammonia-lyases from some species of pseudomonas: taxonomic implication. | 1. histidine ammonia-lyases (histidase ec 4.3.1.3) from pseudomonas testosteroni ncib 10808 and pseudomonas putida ncib 10807 were purified and specific antibody was raised to each separately in a rabbit. 2. immunological cross-reactions of each antibody to histidine ammonia-lyases from various species of pseudomonas were examined by the enzyme inhibition test. 3. the immunological data obtained suggest that these pseudomonas species can be classified into three groups. these cross-reactions ten ... | 1983 | 6862094 |
| partial purification and characterization of a membrane-associated steroid-binding protein from pseudomonas testosteroni. | a steroid-binding protein obtained from the supernatant of the final wash from the preparation of membrane vesicles was purified severalfold to near homogeneity. the protein binds c18 and c19 steroids but has the highest affinity for androstenedione (kd = 1.6 x 10(-10) m). the molecular weight is 51,000 - 58,000. binding activity is slightly inhibited by cu2+, ca2+, and mg2+ and completely inhibited by zn2+. the protein has no detectable steroid degradative activity. analysis of androstenedione ... | 1982 | 6889907 |
| membrane-bound dehydrogenases of pseudomonas testosteroni. | | 1980 | 6931945 |
| membrane-associated steroid-binding proteins of pseudomonas testosteroni. | previous studies have established the presence of periplasmic steroid-binding proteins in induced pseudomonas testosteroni. this study reports the presence of two additional steroid-binding proteins which can be released from the cytoplasmic membrane during preparation of membrane vesicles. these two proteins differ from each other and from the periplasmic protein in some of their characteristics and steroid specificity, and appear to be distinct from the two membrane-bound steroid dehydrogenase ... | 1981 | 6949624 |
| induction of 3-hydroxybenzoate 2-hydroxylase in a pseudomonas testosteroni mutant. | benzoate was established as the inducer of a unique 3-hydroxybenzoate 2-hydroxylase activity found in a pseudomonas testosteroni mutant which is unable to grow on m-hydroxybenzoate as its sole source of carbon and energy. | 1982 | 7054148 |
| testosterone-dependent oxygen consumption in membrane vesicles of pseudomonas testosteroni. | oxygen consumption was measured in membrane vesicles of pseudomonas testosteroni using conditions similar to those identified for testosterone transport in these vesicles. testosterone and nad+, which are primary requirements for testosterone transport, were both required for maximum oxygen consumption suggesting that testosterone transport and oxygen consumption were linked. testosterone-dependent oxygen consumption was inhibited by 95% by 1 mm kcn indicating that the electron-transport chain c ... | 1982 | 7109594 |
| mass spectrometric studies of a modified active-site tetrapeptide from delta 5-3-ketosteroid isomerase of pseudomonas testosteroni. | examination of the product of affinity labeling of delta 5-3-ketosteroid isomerase (ec 5.3.3.1) of pseudomonas testosteroni by the suicide substrate [7-3h]5,10-secoestr-5-yne-3,10,17-trione has demonstrated that the steroid becomes bound by an acid- and base-labile linkage to an active-site peptide representing residues 55-58 (h2n-tyr-ala-asn-ser-co2h) of the primary structure (penning, t. m., and talalay, p. (1981) j. biol. chem. 256, 6851-6858). upon release of the steroid by mild acid hydroly ... | 1982 | 7130168 |
| 2-pyrone-4,6-dicarboxylic acid, a catabolite of gallic acids in pseudomonas species. | 2-pyrone-4,6-dicarboxylate hydrolase was purified from 4-hydroxybenzoate-grown pseudomonas testosteroni. gel filtration and electrophoretic measurements indicated that the preparation was homogeneous and gave a molecular weight of 37,200 for the single subunit of the enzyme. hydrolytic activity was dependent upon a functioning sulfhydryl group(s) and was freely reversible; the equilibrium position was dependent upon ph, with equimolar amounts of pyrone and open-chain form present at ph 7.9. sinc ... | 1982 | 7142106 |
| irreversible inactivation of delta 5-3-ketosteroid isomerase of pseudomonas testosteroni by acetylenic suicide substrates. mechanism of formation and properties of the steroid-enzyme adduct. | | 1981 | 7240247 |
| a reinvestigation of the mechanism of delta 5-3-ketosteroid isomerase from bovine adrenal glands. | the mechanism of the isomerization of androst-5-ene 3,17-dione by the isomerase of bovine adrenals has been reinvestigated using the methodology previously developed for the study of the bacterial enzyme of pseudomonas testosteroni. however, owing to the lower activity of the mammalian enzyme, competitive non-enzymic reaction cannot be neglected. it has been shown that even in the absence of spontaneous isomerization, epimerization and exchange of the label on c4 takes place in the buffer. this ... | 1981 | 7284399 |
| inactivation of delta 5-3-oxo steroid isomerase with active-site-directed acetylenic steroids. | several steroid analogues containing conjugated acetylenic ketone groups as part of a seco-ring structure or as substituents on the intact steroid system are irreversible inhibitors of delta 5-3-oxo steroid isomerase (ec 5.3.3.1) from pseudomonas testosteroni. thus 10 beta-(1-oxoprop-2-ynyl)oestr-4-ene-3,17-dione (i), 5,10-seco-oestr-4-yne-3,10,17-trione (ii), 17 beta-hydroxy-5,10-seco-oestr-4-yne-3,10-dione (iii) and 17 beta-(1-oxoprop-2-ynyl)androst-4-en-3-one (iv) irreversibly inactivate isom ... | 1981 | 7305923 |
| bioconversion of m-hydroxybenzoate to 2,3-dihydroxybenzoate by mutants of pseudomonas testosteroni. | mutans of pseudomonas testosteroni were isolated for their inability to grow on m-hydroxybenzoate as sole carbon source. these mutants hydroxylated m-hydroxybenzoate for form 2,3-dihydroxybenzoate in high yeilds. the bioconversion described in this report represents the first reported example of 3-hydroxybenzoate 2-hydroxylase activity. | 1980 | 7354000 |
| the purification and characterization of delta 5-3-ketosteroid isomerase from pseudomonas putida, a cysteine-containing isomerase. | a delta 5-3-ketosteroid isomerase (ec 5.3.3.1) has been isolated from pseudomonas putida biotype b and purified to homogeneity. this previously undescribed steroid isomerase resembles that isolated from pseudomonas testosteroni (talalay, p., and wang, v.s. (1955) biochim. biophys. acta 18, 300-301). the enzyme is induced by various steroids, has a subunit molecular weight of 13,750 +/- 250, a pi of 4.8 +/- 0.1 has a specific activity of 40,000 units/mg, using 5-androstene-3,17-dione as the subst ... | 1980 | 7358699 |
| on the number of steroid-binding sites of delta 5-3-oxosteroid isomerase. | the number of steroid-binding sites in delta 5-3-oxosteroid isomerase of pseudomonas testosteroni (ec 5.3.3.1) has been determined from measurements of the red shift of the ultraviolet chromophore of 19-nortestosterone upon binding to the enzyme. the experiments include spectroscopic measurements when limiting concentrations of either 19-nortestosterone or isomerase are titrated with varying concentrations of the complementary ligand. analysis of the results indicates one binding site per subuni ... | 1980 | 7371643 |
| an active site phenylalanine of 3-oxo-delta 5-steroid isomerase is catalytically important for proton transfer. | 3-oxo-delta 5-steroid isomerase (ksi) from pseudomonas testosteroni catalyzes the isomerization of a variety of 3-oxo-delta 5-steroids to their conjugated delta 4-isomers through the intermediate formation of a dienolate ion. this dienolate is formed by proton transfer from c-4 of the substrate to asp-38, which then protonates the dienolate at c-6. catalysis is enhanced by electrophilic assistance (hydrogen bonding) to the 3-oxygen by tyr-14. we have investigated the effect of modifying phenylal ... | 1995 | 7492546 |
| cloning, sequencing and expression of pseudomonas testosteroni gene encoding 3 alpha-hydroxysteroid dehydrogenase. | we describe the cloning, sequencing and expression of the 3 alpha-hydroxysteroid dehydrogenase (3 alpha-hsd) gene of pseudomonas testosteroni. a genomic library of p. testosteroni total dna constructed from sauiiia digests ligated to an lambda gt11 vector was probed with a polyclonal antibody raised against purified enzyme. subclones derived from a recombinant phage containing a 1746 bp insert were sequenced and found to contain an open reading frame of 696 bp that corresponds to a protein of 23 ... | 1995 | 7495703 |
| use of a simplified cell blot technique and 16s rrna-directed probes for identification of common environmental isolates. | a simple technique in which rrna-targeted oligodeoxynucleotide probes are used to identify bacteria immobilized on membranes is described. by using colony lifts, bacteria are directly transferred from plates to untreated nitrocellulose membranes. alternatively, cells resuspended from colonies can be applied to membranes by using a vacuum manifold under high-salt conditions. blotted cells are baked and hybridized under stringent conditions by using standard protocols. treatment of blotted cells w ... | 1993 | 7504429 |
| quinoline 2-oxidoreductase and 2-oxo-1,2-dihydroquinoline 5,6-dioxygenase from comamonas testosteroni 63. the first two enzymes in quinoline and 3-methylquinoline degradation. | the enzymes catalysing the first two steps of quinoline and 3-methylquinoline degradation by comamonas testosteroni 63 were investigated. quinoline 2-oxidoreductase, which catalyses the hydroxylation of (3-methyl-)quinoline to (3-methyl-)2-oxo-1,2-dihydroquinoline, was purified to apparent homogeneity. the native enzyme, with a molecular mass of 360 kda, is composed of three non-identical subunits (87, 32, and 22 kda), occurring in a ratio of 1.16:1:0.83. containing fad, molybdenum, iron, and ac ... | 1995 | 7556204 |
| insights into the catalytic mechanism and active-site environment of comamonas testosteroni delta 5-3-ketosteroid isomerase as revealed by site-directed mutagenesis of the catalytic base aspartate-38. | delta 5-3-ketosteroid isomerase (ksi) of comamonas testosteroni catalyzes the isomerization of a wide variety of delta 5(6) and delta 5(10) steroids through the formation of an enzyme bound dienol(ate) intermediate. asp-38 has been strongly implicated in catalysis, apparently serving as a proton shuttle. in this paper the results of a detailed kinetic characterization of the ksi mutants d38e and d38h are presented. both mutants retain significant activity, with kcat and kcat/km values 10(3)-10(4 ... | 1995 | 7578024 |
| purification and characterization of the comamonas testosteroni b-356 biphenyl dioxygenase components. | in this report, we describe some of the characteristics of the comamonas testosteroni b-356 biphenyl (bph)-chlorobiphenyl dioxygenase system, which includes the terminal oxygenase, an iron-sulfur protein (ispbph) made up of an alpha subunit (51 kda) and a beta subunit (22 kda) encoded by bpha and bphe, respectively; a ferredoxin (ferbph; 12 kda) encoded by bphf; and a ferredoxin reductase (redbph; 43 kda) encoded by bphg. ispbph subunits were purified from b-356 cells grown on bph. since highly ... | 1995 | 7592440 |
| molecular cloning, expression in streptomyces lividans, and analysis of a gene cluster from arthrobacter simplex encoding 3-ketosteroid-delta 1-dehydrogenase, 3-ketosteroid-delta 5-isomerase and a hypothetical regulatory protein. | the arthrobacter simplex gene coding for 3-ketosteroid-delta 1-dehydrogenase, a key enzyme in the degradation of the steroid nucleus, was cloned in streptomyces lividans. nucleotide sequence analysis revealed that the gene for 3-ketosteroid-delta 1-dehydrogenase (ksdd) is clustered with at least two more genes possibly involved in steroid metabolism. upstream of ksdd, we found a gene, ksdr, encoding a hypothetical regulatory protein that shows homologies to kdgr, the negative regulator of pectin ... | 1995 | 7596291 |
| quinohaemoprotein ethanol dehydrogenase from comamonas testosteroni. purification, characterization, and reconstitution of the apoenzyme with pyrroloquinoline quinone analogues. | pyrroloquinoline-quinone(pqq)-free quinohaemoprotein ethanol dehydrogenase (qh-edh) apoenzyme was isolated from ethanol-grown comamonas testosteroni. the purified apoenzyme, showing a single band of 71 kda on native gel electrophoresis, could be only partially converted into active holoenzyme by addition of pqq in the presence of calcium ions. in addition to a band with a molecular mass of 71 kda, additional bands of 51 kda and 25 kda were observed with sds/page. analysis of the n-terminal seque ... | 1995 | 7601151 |
| characterization of the interaction between pqq and heme c in the quinohemoprotein ethanol dehydrogenase from comamonas testosteroni. | quinohemoprotein ethanol dehydrogenase from comamonas testosteroni (qh-edh) contains two cofactors, 2,7,9-tricarboxy-1h-pyrrolo[2,3-f]quinoline-4,5-dione (pqq) and heme c. since previous studies on the kinetics of this enzyme suggested that both participate in electron transfer, spectroscopic investigations were performed of the oxidized and reduced holo- and apoenzyme (without pqq but with heme c) to reveal the nature of the interaction between the two redox centers. from this it appears that t ... | 1995 | 7626615 |
| characterization by arbitrary primer polymerase chain reaction of polychlorinated biphenyl (pcb)-degrading strains of comamonas testosteroni isolated from pcb-contaminated soil. | in this study, we isolated and characterized biphenyl (bp) and polychlorinated biphenyl (pcb) degrading bacterial strains found in pcb-contaminated soil from an auto manufacturing plant located in syracuse, new york. twenty-one bp and pcb-degrading bacteria were randomly selected to form a representative sample of the bacterial population present at the site. of the 21 bacteria, 13 were identified as comamonas testosteroni, constituting about 60% of the bacterial population examined. other pcb d ... | 1995 | 7641143 |
| aliphatic nitrilase from a soil-isolated comamonas testosteroni sp.: gene cloning and overexpression, purification and primary structure. | an aliphatic nitrilase, active on adiponitrile and cyanovaleric acid, was identified and purified from comamonas testosteroni sp. (ct). oligodeoxyribonucleotide probes were designed from limited amino acid (aa) sequence information and used to clone the corresponding gene, named nita. high homologies were found at the aa level between ct nitrilase and the sequences of known nitrilases. multi-alignment of sequenced nitrilases suggests that cys163 of ct plays an essential role in the active site. ... | 1995 | 7642130 |
| ultraviolet resonance raman spectroscopy of delta 5-3-ketosteroid isomerase revisited: substrate polarization by active-site residues. | the delta 5-3-ketosteroid isomerase (ec 5.3.3.1) of pseudomonas testosteroni promotes extremely rapid conversion of delta 5- to delta 4-3-ketosteroids by a conservative intramolecular proton transfer via an enolic intermediate. the competitive inhibitor 19-nortestosterone displays marked spectroscopic changes upon binding to the enzyme, but the mechanisms responsible for these changes have not been unequivocally established. ultraviolet resonance raman (uvrr) spectra are reported for 19-nortesto ... | 1995 | 7703258 |
| identification of active site residues by site-directed mutagenesis of delta 5-3-ketosteroid isomerase from pseudomonas putida biotype b. | in order to assess the roles of specific amino acid residues in the delta 5-3-ketosteroid isomerase from pseudomonas putida biotype b during catalysis, we replaced aspartic acid 40 with asparagine (d40n) and tyrosine 16 with phenylalanine (y16f) in the enzyme by site-directed mutagenesis. both purified mutant enzymes resulted in profound decreases in catalytic activities, 10(3.3)-fold in the y16f mutant and 10(6.2)-fold in the d40n mutant. aspartic acid 40 and tyrosine 16 of the enzyme are the c ... | 1995 | 7730300 |
| sequence of the bphd gene encoding 2-hydroxy-6-oxo-(phenyl/chlorophenyl)hexa-2,4-dienoic acid (hop/cpda) hydrolase involved in the biphenyl/polychlorinated biphenyl degradation pathway in comamonas testosteroni: evidence suggesting involvement of ser112 in catalytic activity. | the nucleotide sequence of bphd, encoding 2-hydroxy-6-oxo-(phenyl/chlorophenyl)hexa-2,4-dienoic acid hydrolase involved in the biphenyl/polychlorinated biphenyl degradation pathway of comamonas testosteroni strain b-356, was determined. comparison of the deduced amino-acid sequence with published sequences led to the identification of a 'lipase box', containing a consensus pentapeptide sequence glyxaaserxaagly. this suggested that the mechanism of action of this enzyme may involve an asp-ser-his ... | 1995 | 7737519 |
| ultraviolet spectroscopic evidence for decreased motion of the active site tyrosine residue of delta 5-3-ketosteroid isomerase by steroid binding. | delta 5-3-ketosteroid isomerase (ec 5.3.3.1) from pseudomonas testosteroni catalyzes the highly efficient conversion of delta 5-3-ketosteroids to delta 4-3-ketosteroids by a stereoselective and intramolecular transfer of the 4 beta-proton to the 6 beta-position. tyr-14 is the critical general acid and asp-38 is the general base involved in catalysis. the uv absorption bandwidths of tyr-14 were much narrower than those of the other two tyrosines (tyr-55 and tyr-88) of isomerase or of the n-acetyl ... | 1995 | 7756287 |
| purification and properties of bile acid sulfate sulfatase from pseudomonas testosteroni. | the bile acid sulfate sulfatase (bss) produced by pseudomonas testosteroni was purified and characterized. chromatofocusing behavior and amino acid sequence over twelve amino acid residues from n-terminus of the enzyme indicated that bss was composed of two isoforms of which molecular weights were 125,000 and 103,000. each isoform was a homodimer of a subunit of which molecular weight was 53,000 or 51,000, respectively. the optimum ph was 8.5 and bss was stable at ph 5.8-8.0. the thermostability ... | 1994 | 7764976 |
| enzymatic and nonenzymatic polarizations of alpha,beta-unsaturated ketosteroids and phenolic steroids. implications for the roles of hydrogen bonding in the catalytic mechanism of delta 5-3-ketosteroid isomerase. | ketosteroids (e.g., 19-nortestosterone) and phenolic steroids (e.g., 17 beta-estradiol and 17 beta-dihydroequilenin), which are potent competitive inhibitors of delta 5-3-ketosteroid isomerase (isomerase, ec 5.3.3.1) of pseudomonas testosteroni, undergo significant polarization upon binding to the active site of the enzyme. the 10 nm red shift of the uv absorption maximum of the enone chromophore of 19-nortestosterone, which occurs in the enzyme-steroid complex, resembles that observed when this ... | 1995 | 7819234 |
| regulation of the hema gene during 5-aminolevulinic acid formation in pseudomonas aeruginosa. | the general tetrapyrrole precursor 5-aminolevulinic acid is formed in bacteria via two different biosynthetic pathways. members of the alpha group of the proteobacteria use 5-aminolevulinic acid synthase for the condensation of succinyl-coenzyme a and glycine, while other bacteria utilize a two-step pathway from aminoacylated trna(glu). the trna-dependent pathway, involving the enzymes glutamyl-trna reductase (encoded by hema) and glutamate-1-semialdehyde-2,1-aminomutase (encoded by heml), was d ... | 1995 | 7883699 |
| improved purification of steroid 1:2-dehydrogenase from nocardia opaca and partial characterization of its cloned gene sequence. | we have purified a steroid-inducible 1:2-dehydrogenase from nocardia opaca. the final enzyme preparation was purified 120-fold with a recovery of 38%. the n-terminal amino acid sequence was determined to be: met-gln-asp-trp-thr-ser-glu-(cys)-asp-val-leu-val-val-gly-. from the genomic library of nocardia opaca in the plasmid puc19, a clone designated as pstd23 containing a 0.9 kb kpni-psti fragment was found to hybridize with an oligonucleotide probe corresponding to the first six amino acids fro ... | 1993 | 7916596 |
| identification and mapping of the gene translation products involved in the first steps of the comamonas testosteroni b-356 biphenyl/chlorobiphenyl biodegradation pathway. | in this study, we have mapped comamonas testosteroni b-356 genes encoding enzymes for the conversion of biphenyl and 4-chlorobiphenyl into the corresponding meta-cleavage compounds onto a 6.3-kb dna fragment, and we have determined the subunit composition of the enzymes involved in this pathway. the various proteins encoded by this 6.3-kb dna fragment and by subclones derived from it were overexpressed and selectively labelled using the t7 polymerase promoter system in escherichia coli. they wer ... | 1994 | 7954108 |
| terephthalate 1,2-dioxygenase system from comamonas testosteroni t-2: purification and some properties of the oxygenase component. | comamonas testosteroni t-2, grown in terephthalate (ter)-salts medium, synthesizes inducible enzymes that convert ter to (1r,2s)-dihydroxy-3,5-cyclohexadiene-1,4-dicarboxylic acid (dcd) and protocatechuate (pc). anion-exchange chromatography of cell extracts yielded two sets of fractions, r and z, that were necessary for oxygenation of ter to dcd; we termed this activity the ter dioxygenase system (terdos). an nad(+)-dependent dcd dehydrogenase, which converted dcd to pc, overlapped all fraction ... | 1994 | 7961417 |
| description of the kinetic mechanism and the enantioselectivity of quinohaemoprotein ethanol dehydrogenase from comamonas testosteroni in the oxidation of alcohols and aldehydes. | initial rate studies were performed on the oxidation of (racemic) alcohols as well as aldehydes by quinohaemoprotein ethanol dehydrogenase, type 1, from comamonas testosteroni with potassium ferricyanide as electron acceptor. the data could be fitted with an equation derived for a mechanism (hexa-uni ping-pong) in which alcohols are oxidized to the corresponding carboxylic acids and the intermediate aldehyde becomes released from the enzyme. however, for some substrates it was necessary to assum ... | 1994 | 8001568 |
| characterization of the three tyrosine residues of delta 5-3-ketosteroid isomerase by time-resolved fluorescence and circular dichroism. | delta 5-3-ketosteroid isomerase (ec 5.3.3.1) of pseudomonas testosteroni converts delta 5-3-ketosteroids to delta 4-3-ketosteroids via an enolic intermediate. site-specific mutagenesis has identified tyr-14 and asp-38 as the catalytically essential general acid and base, respectively. three tyrosine residues (tyr-14, tyr-55, and tyr-88) are the only significant fluorophores in the wild-type isomerase. recent studies of the steady-state fluorescence of the wild-type enzyme and all six mutant enzy ... | 1994 | 8003507 |
| regulation of chloro- and methylphenol degradation in comamonas testosteroni jh5. | comamonas testosteroni jh5 was isolated from a mixed bacterial culture enriched on different chloro- and methylphenols. the strain completely mineralized a mixture consisting of 4-chlorophenol (4-cp) and 4-methylphenol (4-mp). during degradation of the mixture, 4-hydroxybenzyl alcohol, 4-hydroxybenzaldehyde, 4-hydroxybenzoic acid, and 4-chlorocatechol were detected as short-lived intermediates. mineralization of 4-cp and that of 4-mp occurred successively and were accompanied by diauxic growth, ... | 1994 | 8074514 |
| kinetics and mechanism of heterogeneous hydrolysis of poly[(r)-3-hydroxybutyrate] film by pha depolymerases. | the kinetics and mechanism of enzymatic degradation on the surface of poly[(r)-3-hydroxybutyrate] (p[(r)-3hb]) film have been studied using three types of extracellular poly(hydroxyalkanoate) (pha) depolymerases from alcaligenes faecalis, pseudomonas picketti and comamonas testosteroni. the monomer and dimer of 3-hydroxybutyric acid were produced during the course of the enzymatic degradation of p[(r)-3hb] film, and the rate of production was determined by monitoring the increase in absorbance a ... | 1993 | 8110658 |
| extent of proton transfer in the transition states of the reaction catalyzed by the delta 5-3-ketosteroid isomerase of comamonas (pseudomonas) testosteroni: site-specific replacement of the active site base, aspartate 38, by the weaker base alanine-3-sulfinate. | previous studies of the mechanism of the steroid isomerase of comamonas (pseudomonas) testosteroni have identified aspartate 38 as the proton porter which transfers the substrate's 4 beta proton to the 6 beta position of the product. consequently, aspartate 38 functions as a base in the deprotonation of the substrate to form a dienol or dienolate intermediate, which then undergoes reprotonation from protonated aspartate 38 at c-6 beta to give the product. we have tried to characterize the transi ... | 1994 | 8117731 |
| mechanism of the reaction catalyzed by delta 5-3-ketosteroid isomerase of comamonas (pseudomonas) testosteroni: kinetic properties of a modified enzyme in which tyrosine 14 is replaced by 3-fluorotyrosine. | tyrosine 14 of delta 5-3-ketosteroid isomerase plays an important role in the function of the enzyme, since its replacement by phenylalanine results in a decrease in kcat by a factor of 10(-4.7). this result and the fact that this residue resides in the enzyme's substrate binding site and is in close proximity to c-2 of the bound steroid suggests that it functions as an electrophile in the catalytic mechanism by protonation of or hydrogen bonding to the c-3 carbonyl oxygen of the substrate. in o ... | 1994 | 8117732 |
| factors affecting pcb degradation by an implanted bacterial strain in soil microcosms. | pseudomonas testosteroni b-356 was able to degrade approximately 50% of the aroclor 1242 mixture in shaken culture. the aims of the present study were to evaluate the capabilities of this bacterial strain to degrade pcbs in soil microcosms and to identify some of the factors likely to favor the degradative performance of the implanted bacteria. the presence of biphenyl as cosubstrate was the most important factor affecting pcb degradation in soil. however, because biphenyl was rapidly depleted i ... | 1993 | 8358671 |