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A M Cook

Publications and source records attributed to A M Cook.

At least 37 records · Page 2Linked to original sources
PubMed1998

Methods for studying leukocyte filterability in undiluted blood from intermittent claudicants.

Leukocytes are characterised from their influence on the filterability of undiluted blood from patients with PAOD (intermittent claudicants - Fontaine Stage II) and a group of sex and age-matched controls. Undiluted blood was filtered through 5 microm Nuclepore (Hemafil) filters for 300 s, at 711 Pa and room temperature, using a custom-made constant pressure filtrometer. Four populations of leukocytes are identified in both groups. In the control group, 94.4% of the leukocytes are identified as fast leukocytes with a transit time of 1.8 s. The remaining white cells are recognised as slow flowing leukocytes and subdivided into three further sub-populations. The first of these (SL1; 2.8% of total leukocyte count) is characterised by a transit time of 31.7 s, a second population (SL2; 1.5% of total leukocyte count) by a transit time of 145.8 s while the remaining cells are identified as pore blockers (PB) under these conditions. A similar rheological classification is valid in the patients but the sum of the three minor populations is elevated compared to controls (p = 0.001) although there is no overall leukocytosis. The only significant difference in flow properties of any blood cells, between the two groups of volunteers, is seen in the major population of leukocytes with an elevated transit time of 2.4 s. Stepwise regression analysis identifies the concentration of fast leukocytes, SL2 and PB as the major variables affecting blood flow through the filter. It is argued that the higher concentration of SL2 and PB probably reflect the increased sensitivity of neutrophils to physical stimuli.

Aged↗
PubMed1997

Thiosulfate as a metabolic product: the bacterial fermentation of taurine.

Thiosulfate (S2O32-) is a natural product that is widely utilized in natural ecosystems as an electron sink or as an electron donor. However, the major biological source(s) of this thiosulfate is unknown. We present the first report that taurine (2-aminoethanesulfonate), the major mammalian solute, is subject to fermentation. This bacterial fermentation was found to be catalyzed by a new isolate, strain GKNTAU, a strictly anaerobic, gram-positive, motile rod that formed subterminal spores. Thiosulfate was a quantitative fermentation product. The other fermentation products were ammonia and acetate, and all could be formed by cell-free extracts.

Acetates↗
PubMed1997

Taurine reduction in anaerobic respiration of Bilophila wadsworthia RZATAU.

Organosulfonates are important natural and man-made compounds, but until recently (T. J. Lie, T. Pitta, E. R. Leadbetter, W. Godchaux III, and J. R. Leadbetter. Arch. Microbiol. 166:204-210, 1996), they were not believed to be dissimilated under anoxic conditions. We also chose to test whether alkane- and arenesulfonates could serve as electron sinks in respiratory metabolism. We generated 60 anoxic enrichment cultures in mineral salts medium which included several potential electron donors and a single organic sulfonate as an electron sink, and we used material from anaerobic digestors in communal sewage works as inocula. None of the four aromatic sulfonates, the three unsubstituted alkanesulfonates, or the N-sulfonate tested gave positive enrichment cultures requiring both the electron donor and electron sink for growth. Nine cultures utilizing the natural products taurine, cysteate, or isethionate were considered positive for growth, and all formed sulfide. Two clearly different pure cultures were examined. Putative Desulfovibrio sp. strain RZACYSA, with lactate as the electron donor, utilized sulfate, aminomethanesulfonate, taurine, isethionate, and cysteate, converting the latter to ammonia, acetate, and sulfide. Strain RZATAU was identified by 16S rDNA analysis as Bilophila wadsworthia. In the presence of, e.g., formate as the electron donor, it utilized, e.g., cysteate and isethionate and converted taurine quantitatively to cell material and products identified as ammonia, acetate, and sulfide. Sulfite and thiosulfate, but not sulfate, were utilized as electron sinks, as was nitrate, when lactate was provided as the electron donor and carbon source. A growth requirement for 1,4-naphthoquinone indicates a menaquinone electron carrier, and the presence of cytochrome c supports the presence of an electron transport chain. Pyruvate-dependent disappearance of taurine from cell extracts, as well as formation of alanine and release of ammonia and acetate, was detected. We suspected that sulfite is an intermediate, and we detected desulfoviridin (sulfite reductase). We thus believe that sulfonate reduction is one aspect of a respiratory system transferring electrons from, e.g., formate to sulfite reductase via an electron transport system which presumably generates a proton gradient across the cell membrane.

Acetates↗
PubMed1997

Conjugative plasmids and the degradation of arylsulfonates in Comamonas testosteroni.

Comamonas testosteroni T-2 degrades p-toluenesulfonate (TSA) via p-sulfobenzoate (PSB) and protocatechuate and degrades toluenecarboxylate via terephthalate (TER) and protocatechuate. The appropriate genes are expressed in at least five regulatory units, some of which are also found in C. testosteroni PSB-4 (F. Junker, R. Kiewitz, and A. M. Cook, J. Bacteriol. 179:919-927, 1997). C. testosteroni T-2 was found to contain two plasmids, pTSA (85 kbp) and pT2T (50 kbp); a TSA- mutant (strain TER-1) contained only plasmid pT2T. C. testosteroni PSB-4, which does not degrade TSA, contained one plasmid, pPSB (85 kbp). The type strain contained no plasmids. Conjugation experiments showed that plasmid pTSA (possibly in conjunction with pT2T) was conjugative, and the single copy of the TSA operon (tsaMBCD) with its putative regulator gene (tsaR) in strain T-2 was found on plasmid pTSA, which also carried the PSB genes (psbAC) and presumably transport for both substrates. Plasmid pTSA was assigned to the IncP1 beta group and was found to carry two copies of insertion element IS1071. Plasmid pPSB (of strain PSB-4), which could be maintained in strains with plasmid pTSA or pT2T, was also conjugative and was found to carry the PSB genes as well as to contain two copies of IS1071. In attempted conjugations with the type strain, no plasmid was recovered, but the PSB+ transconjugant carried two copies of IS1071 in the chromosome. We presume the PSB genes to be located in a composite transposon. The genes encoding the putative TER operon and degradation of protocatechuate, with the meta cleavage pathway, were attributed a chromosomal location in strains T-2 and PSB-4.

Amino Acid Sequence↗
PubMed1997

Characterization of the p-toluenesulfonate operon tsaMBCD and tsaR in Comamonas testosteroni T-2.

Comamonas testosteroni T-2 uses a standard, if seldom examined, attack on an aromatic compound and oxygenates the side chain of p-toluenesulfonate (TS) (or p-toluenecarboxylate) to p-sulfobenzoate (or terephthalate) prior to complete oxidation. The expression of the first three catabolic enzymes in the pathway, the TS methyl-monooxygenase system (comprising reductase B and oxygenase M; TsaMB), p-sulfobenzyl alcohol dehydrogenase (TsaC), and p-sulfobenzaldehyde dehydrogenase (TsaD), is coregulated as regulatory unit R1 (H. R. Schlafli Oppenberg, G. Chen, T. Leisinger, and A. M. Cook, Microbiology [Reading] 141:1891-1899, 1995). The components of the oxygenase system were repurified, and the N-terminal amino acid sequences were confirmed and extended. An internal sequence of TsaM was obtained, and the identity of the [2Fe-2S] Rieske center was confirmed by electron paramagnetic resonance spectroscopy. We purified both dehydrogenases (TsaC and TsaD) and determined their molecular weights and N-terminal amino acid sequences. Oligonucleotides derived from the partial sequences of TsaM were used to identify cloned DNA from strain T-2, and about 6 kb of contiguous cloned DNA was sequenced. Regulatory unit R1 was presumed to represent a four-gene operon (tsaMBCD) which was regulated by the LysR-type regulator, TsaR, encoded by a deduced one-gene transcriptional unit. The genes for the inducible TS transport system were not at this locus. The oxygenase system was confirmed to be a class IA mononuclear iron oxygenase, and class IA can now be seen to have two evolutionary groups, the monooxygenases and the dioxygenases, though the divergence is limited to the oxygenase components. The alcohol dehydrogenase TsaC was confirmed to belong to the short-chain, zinc-independent dehydrogenases, and the aldehyde dehydrogenase TsaD was found to resemble several other aldehyde dehydrogenases. The operon and its putative regulator are compared with units of the TOL plasmid.

Alcohol Dehydrogenase↗
PubMed1996

Comparison of mitozantrone and epirubicin in advanced breast cancer.

Forty chemotherapy naive patients with metastatic or locally advanced breast cancer were treated in a randomized trial comparing mitozantrone 14 mg/m2 with epirubicin 75 mg/m2 given intravenously at 3-weekly intervals. There was a 40% (95% confidence interval (CI) 8-72; P = 0.013) higher partial response rate with epirubicin (11/18) than with mitozantrone (4/19). Epirubicin caused significantly more alopecia (difference 76%; 95% CI 57-96; P < 0.0001) and nausea/vomiting (difference = 38%; 95% CI 10-67; P = 0.01). Three patients who received long courses of epirubicin experienced cardiac failure; two were proved to have cardiomyopathy. The median survival for the epirubicin and mitozantrone groups were 9.5 and 8 months respectively. Thus, although epirubicin gave a higher response rate it also caused more toxicity.

Antineoplastic Agents↗
PubMed1996

Degradative pathways for p-toluenecarboxylate and p-toluenesulfonate and their multicomponent oxygenases in Comamonas testosteroni strains PSB-4 and T-2.

Three multicomponent oxygenases involved in the degradation of p-toluenesulfonate and p-toluenecarboxylate and the regulation of their synthesis have been examined in three strains (T-2, PSB-4 and TER-1) of Comamonas testosteroni. Strain T-2 utilizes p-toluenesulfonate as a source of carbon and energy for growth via p-sulfobenzoate and protocatechuate, and p-toluenecarboxylate via terephthalate and protocatechuate, and has the unusual property of requiring the reductase (TsaB) of the toluenesulfonate methyl monooxygenase system (TsaMB) in an incompletely expressed sulfobenzoate dioxygenase system (PsbAC) [Schläfli Oppenberg, H.R., Chen, G., Leisinger, T. & Cook, A. M. (1995). Microbiology 141, 1891-1899]. The independently isolated C. testosteroni PSB-4 utilized only sulfobenzoate and terephthalate via protocatechuate. Mutant TER-1, derived from strain T-2, utilized only terephthalate via protocatechuate. We detected no enzymes of the pathway from toluenesulfonate to sulfobenzoate in strains PSB-4 and TER-1, and confirmed by PCR and Southern blot analysis that the genes (tsaMB) encoding toluenesulfonate monooxygenase were absent. We concluded that, in strain PSB-4, the regulatory unit encoding the genes for the conversion of toluenesulfonate to sulfobenzoate was missing, and that generation of mutant TER-1 involved deletion of this regulatory unit and of the regulatory unit encoding desulfonation of sulfobenzoate. The degradation of sulfobenzoate in strain PSB-4 was catalysed by a fully inducible sulfobenzoate dioxygenase system (PsbACPSB-4), which, after purification of the oxygenase component (PsbAPSB-4), turned out to be indistinguishable from the corresponding component from strain T-2 (PsbAT-2). Reductase PsbCPSB-4, which we could separate but not purify, was active with oxygenase PsbAPSB-4 and PsbAT-2. Oxygenase PsbAPSB-4 was shown by electron paramagnetic resonance spectroscopy to contain a Rieske [2Fe-2S] centre. The enzyme system oxygenating terephthalate was examined and the oxygenase component purified and characterized. The oxygenase component in strains T-2 (and mutant TER-1) and PSB-4 were indistinguishable. The reductase component, which we separated but failed to purify, was active with the oxygenase from all strains. Gains and losses of blocks of genes in evolution is discussed.

Benzenesulfonates↗
PubMed1996

The assimilation of sulfur from multiple sources and its correlation with expression of the sulfate-starvation-induced stimulon in Pseudomonas putida S-313.

Conditions were optimized for the batch growth of Pseudomonas putida S-313 under sulfur-limited conditions. P. putida grew exponentially with sulfate as the sole source of sulfur, and growth was concomitant with the utilization of sulfate until it was exhausted. A further 20% of protein was synthesized after the apparent disappearance of sulfate. A mass balance for the utilized sulfate in cell material was calculated, given the observed molar growth yield of about 3.6 kg protein (mol S)-1 and a sulfur content of 0.41% S in dry matter. Similar data were obtained for growth with cysteine and thiocyanate. The organism also grew exponentially with 4-toluenesulfonate (TS) as sulfur source, essentially as observed with sulfate, except that negligible protein formation after exhaustion of TS was observed. Similar data were also obtained with 4-nitrocatecholsulfate (NCS) and ethanesulfonate. Any substrate pair selected from sulfate, cysteine and thiocyanate was utilized simultaneously, and although one of the pair of substrates was always preferred, growth continued at the same rate when only one substrate remained. Growth after substrate exhaustion was observed. Any substrate pair selected from TS, NCS and ethanesulfonate gave similar data, but with less growth after exhaustion of the sulfur sources. If a mixed substrate pair was chosen from the two groups, the sulfur source from the first-named group was initially used exclusively, and the second source of sulfur was utilized subsequently, after a lag phase. The data are considered to reflect the control of scavenging for sulfur and of distribution of sulfur in the cell exerted by the sulfate-starvation-induced stimulon.

Culture Media↗
PubMed1995

Purification and characterization of the arylsulfatase synthesized by Pseudomonas aeruginosa PAO during growth in sulfate-free medium and cloning of the arylsulfatase gene (atsA).

An arylsulfatase (EC 3.1.6.1) was extracted from Pseudomonas aeruginosa PAO1 and purified 2700-fold to homogeneity. Synthesis of this enzyme was repressed when sulfate, cysteine or thiocyanate was supplied as the sole sulfur source for growth, but derepressed with all other sulfur sources tested. The apparent molecular mass was determined by SDS/PAGE to be 57 kDa, and the enzyme was presumed to be a monomer after gel filtration chromatography. The arylsulfatase showed maximal activity at 57 degrees C and pH 8.9, and a Km of 105 microM for 4-nitrocatecholsulfate. Despite previous reports that both inducible and derepressible forms of arylsulfatase exist in P. aeruginosa, we found only one enzyme under a variety of growth conditions: a sulfate-repressed enzyme with a native isoelectric point of 4.76. The gene encoding this enzyme (atsA) was isolated by complementation of a Tn5-751 mutant of P. aeruginosa PAO1. Sequencing revealed a 1602-bp reading frame encoding a 534-amino-acid protein with sequence similarity to known bacterial and eukaryotic arylsulfatases (30-40% and 25-30% identity, respectively), but lacking the signal peptide which is present in all known sequences. The lack of this signal peptide suggests that the P. aeruginosa arylsulfatase is neither periplasmic nor membrane-associated, unlike other known arylsulfatases. The atsA gene was located at 15-17' on the P. aeruginosa genome by Southern hybridization. Only a single copy was observed under moderate stringency conditions.

Amino Acid Sequence↗
PubMed1995

Dynamic gradient-echo and fat-suppressed spin-echo contrast-enhanced MRI of the breast.

PURPOSE: To evaluate heavily T2-weighted, dynamic contrast-enhanced and fat-suppressed magnetic resonance imaging (MRI) of the breast in comparison with conventional imaging and fine needle aspiration cytology (FNAC). PATIENTS AND METHODS: Fifty patients with surgically/pathologically proven breast disease were examined pre-operatively by MRI. The majority, 45 patients, had invasive carcinoma. T1-weighted spin-echo, T2-weighted fast spin-echo (with chemical-shift-selective fat-suppression in 20 cases), rapid dynamic contrast-enhanced gradient-echo and post-contrast fat-suppressed T1-weighted images were obtained. Signal intensity changes during dynamic scanning were assessed qualitatively and quantitatively. Comparison was made with the results of X-ray mammography, ultrasound and fine needle aspiration cytology. RESULTS: Unenhanced MRI was inadequate for determining the location, extent or nature of most lesions even when fat-suppressed T2-weighted images were obtained. Following contrast injection, there was significantly greater enhancement of invasive carcinomas than normal parenchyma. Invasive carcinomas of ductal and lobular subtypes did not differ significantly in their enhancement profiles. Prominent enhancement of the lesion periphery, which was a feature in 33 out of 50 cases (the majority of which were invasive carcinomas) was not due to central tumour necrosis. In four cases, invasive carcinomas which were clearly visible on early dynamic scans could not be identified on post-contrast fat-suppressed images. Lesions that were more numerous or extensive than had been recognised clinically or mammographically were revealed by MRI in 14 patients, though MRI was no more specific than conventional assessment. Invasion of the chest wall was accurately predicted by MRI in three cases. There was excellent correlation between tumour size shown by MRI and histopathology. FNAC was accurate in predicting the final histological diagnosis except in those cases where samples were unsatisfactory. CONCLUSIONS: Contrast-enhanced MRI appears useful in the assessment of suspected malignant breast masses, especially in younger women with predominantly glandular breast tissue. Specificity is no better than FNAC but tumour extent and multifocality are more accurately disclosed than by conventional imaging techniques. Contrast-enhanced chemical-shift-selective fat-suppressed images are sub-optimal in a small proportion of cases (particularly where lesions are located posteriorly) and some benign breast disease may appear misleadingly conspicuous on such images. Morphological features and quantification of lesion enhancement during dynamic scanning presently offer only limited prospects for discrimination between various pathologies. Heavily T2-weighted sequences appear of marginal value.

Adolescent↗
PubMed1995

Stereospecificity of hydride removal from NADH by reductases of multicomponent nonheme iron oxygenase systems.

The stereospecificity of hydride removal from the 4 position of the pyridine ring of NADH by reductases from all three classes of multicomponent nonheme iron oxygenases was examined. The class I and II reductases, modules of which show significant sequence similarity with and which belong to the ferredoxin-NADP+ reductase family of flavin-dependent oxidoreductases, transferred the pro-R hydrogen. By contrast, the class II enzymes, which do not show significant sequence similarity to the class I and III enzymes but modules of which belong to the glutathione reductase family of flavoenzymes, transferred the pro-S hydrogen.

Dioxygenases↗
PubMed1994

Oxygenation and spontaneous deamination of 2-aminobenzenesulphonic acid in Alcaligenes sp. strain O-1 with subsequent meta ring cleavage and spontaneous desulphonation to 2-hydroxymuconic acid.

2-Aminobenzenesulphonic acid (2AS) is degraded by Alcaligenes sp. strain O-1 via a previously detected but unidentified intermediate. A mutant of strain O-1 was found to excrete this intermediate, which was isolated and identified by m.s., 1H- and 13C-n.m.r. as 3-sulphocatechol (3SC). Proteins from cell extracts of strain O-1 were separated by anion-exchange chromatography. A multicomponent oxygenase was observed to convert 1 mol each of NADH, O2 and 2AS into 1 mol each of 3SC, NH3 and NAD+. The enzyme presumably catalysed formation of the ring of a 2-amino-2,3-diol moiety, and elimination in the amino group led to a rearomatization. 3SC was further degraded via meta ring cleavage, which could be prevented by inactivation of the 3-sulphocatechol-2,3-dioxygenase (3SC23O) with 3-chlorocatechol. In Tris buffer, the separated 3SC23O catalysed the reaction of 1 mol each of 3SC and O2 involving a transient yellow intermediate, and release of 1 mol of sulphite and two organic products. The major product was identified by n.m.r. and by g.c./m.s. as 5-carboxypenta-2,4-dien-5-olide (CPDO), an indicator of formation of 2-hydroxymuconic acid (2HM). The second product was identified as the Z,E isomer of 2HM by comparison with authentic material. When the CPDO in the product mixture was chemically hydrolysed to (Z,E)-2HM, 1 mol of (Z,E)-2HM/mol of 3SC was observed. If oxygenation of 3SC by 3SC23O was carried out in phosphate buffer, only a single product was detected, a keto form of 2HM. This dioate was also formed from authentic (Z,E)-2HM in phosphate buffer. Formation of the natural product (Z,E)-2HM from the xenobiotic, 3SC, seems to involve oxygenation to the unstable 2-hydroxy-6-sulphonomuconic acid semialdehyde, which hydrolyses spontaneously to 2HM. There would appear to be at least one spontaneous reaction per enzyme reaction in this pathway.

Alcaligenes↗
PubMed1994

3-Sulphocatechol 2,3-dioxygenase and other dioxygenases (EC 1.13.11.2 and EC 1.14.12.-) in the degradative pathways of 2-aminobenzenesulphonic, benzenesulphonic and 4-toluenesulphonic acids in Alcaligenes sp. strain O-1.

Alcaligenes sp. strain O-1 utilizes three sulphonated aromatic compounds as sole sources of carbon and energy for growth in minimal salts medium-benzenesulphonate (BS), 4-toluenesulphonate (TS) and 2-aminobenzenesulphonate (2AS). The degradative pathway(s) in 2AS-grown cells are initiated with membrane transport, NADH-dependent dioxygenation and meta ring cleavage. The specific activity of the NADH-dependent dioxygenation(s) varied with the growth phase and was maximal near the end of exponential growth for each growth substrate. Cells were harvested at this point from BS-, TS- and 2AS-salts medium. Cells grown with each sulphonated substrate could oxygenate all three compounds, but only 2AS-grown cells consumed 2 mol O2 per mol 2AS or BS or TS. BS- and TS-grown cells consumed 2 mol O2 per mol BS or TS but failed to oxygenate the product of oxygenation of 2AS, 3-sulphocatechol (3SC). These observations were repeated with cell extracts and we concluded that there were two sets of desulphonative pathways in the organism, one for 2AS and one for BS and TS. We confirmed this hypothesis by separating the degradative enzymes from 2AS-, BS- or TS-grown cells. A 2AS dioxygenase system and a 3SC-2,3-dioxygenase (3SC23O) were detected in 2AS-grown cells only. In both BS- and TS-grown cells a dioxygenase system for BS and TS was observed as well as a principal catechol 2,3-dioxygenase (C23O-III), neither of which was present in 2AS-grown cells. The 3SC23O was purified to near homogeneity, found to be monomeric (M(r) 42,000), and to catalyse 2,3-dioxygenation to a product that decayed spontaneously to sulphite and 2-hydroxymuconate. The 2AS dioxygenase system could cause not only deamination of 2AS but also desulphonation of BS and TS. The BS dioxygenase could desulphonate BS and apparently either desulphonate or deaminate 2AS. Strain O-1 thus seems to contain two putative, independently regulated operons involving oxygenation and spontaneous desulphonation(s). One operon encodes at least the 2AS dioxygenase system and 3SC23O whereas the other encodes at least the BS/TS dioxygenase system and C23O-III.

Alcaligenes↗
PubMed1994

Microbial metabolism of sulfur- and phosphorus-containing xenobiotics.

The enzymes involved in the microbial metabolism of many important phosphorus- or sulfur-containing xenobiotics, including organophosphate insecticides and precursors to organosulfate and organosulfonate detergents and dyestuffs have been characterized. In several instances their genes have been cloned and analysed. For phosphonate xenobiotics, the enzyme system responsible for the cleavage of the carbon-phosphorus bond has not yet been observed in vitro, though much is understood on a genetic level about phosphonate degradation. Phosphonate metabolism is regulated as part of the Pho regulon, under phosphate starvation control. For organophosphorothionate pesticides the situation is not so clear, and the mode of regulation appears to depend on whether the compounds are utilized to provide phosphorus, carbon or sulfur for cell growth. The same is true for organosulfonate metabolism, where different (and differently regulated) enzymatic pathways are involved in the utilization of sulfonates as carbon and as sulfur sources, respectively. Observations at the protein level in a number of bacteria suggest that a regulatory system is present which responds to sulfate limitation and controls the synthesis of proteins involved in providing sulfur to the cell and which may reveal analogies between the regulation of phosphorus and sulfur metabolism.

Bacteria↗
PubMed1994

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 fractions R. No significant purification from fraction R, which contained an NADH-dependent reductase function(s) of TERDOS, was attained. Fraction Z, at the end of the gradient, contained essentially one protein, which was further purified by hydrophobic interaction chromatography. This component, Z, had the UV-visible spectrum and electron paramagnetic resonance characteristics of a Rieske [2Fe-2S] protein and was considered to be the oxygenase. M(r)s of about 126,000 for oxygenase Z under native conditions were observed. Oxygenase Z consisted of two subunits, alpha and beta, with M(r)s of 49,000 and 18,000, respectively, under denaturing conditions. We presume that this oxygenase has an alpha 2 beta 2 structure. The sequences of the N-terminal amino acids of each subunit were determined. The activity of the purified enzyme was enhanced about fivefold by addition of Fe2+. In the presence of O2, NADH, and fraction R, component Z catalyzed the stoichiometric transformation of TER to PC, with the intermediate formation of DCD. The reaction was confirmed as a dioxygenation when we observed incorporation of two oxygen atoms from 18O2 into PC. The substrate range of TERDOS appeared to be narrow; apart from TER, only 2,5-dicarboxypyridine and 1,4-dicarboxynaphthalene (of 11 compounds tested) were converted to a product.

Biodegradation, Environmental↗
PubMed1994

Reproducibility of bone ages when performed by radiology registrars: an audit of Tanner and Whitehouse II versus Greulich and Pyle methods.

This audit analysed the Tanner and Whitehouse II twenty bone (TW2) method of bone age assessment which was used in our department, and compared it with the Greulich and Pyle (GP) method. 50 previous bone ages were independently re-calculated by each of three registrars using both techniques, with the time taken to perform each assessment being recorded. For each method the interobserver variation was analysed in terms of the spread of results. The intraobserver variation in TW2 was determined by comparing the bone age originally reported with that subsequently calculated on the same film by the same registrar. The average spread of results was 0.74 years for TW2 method, and 0.96 years for the GP method and this difference is not statistically significant at the 5% level. The average intraobserver variation to TW2 was 0.33 years, but with 95% confidence limits of -0.87 to +1.53 years. The average time taken was 7.9 min for TW2 and 1.4 min for GP assessments. It was concluded that the GP method gave similar reproducibility and was faster than the TW2 method. Following clinical discussion the routine departmental bone age assessment method was changed from the TW2 to the GP method.

Age Determination by Skeleton↗