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Biomedical subjects

B Green

Publications and source records attributed to B Green.

At least 109 records · Page 6Linked to original sources
PubMed1993

Comparison of the proteoglycanolytic activities of human leukocyte elastase and human cathepsin G in vitro and in vivo.

In this study, we evaluated the in vitro and in vivo potency of human leukocyte elastase (HLE) and human cathepsin G (HCG) as proteoglycanases. In vitro evaluation was done using bovine nasal septum aggrecan and aggrecan/hyaluronan aggregate as substrates. Enzyme activity was assessed by the ability of the proteinases to abrogate the ability of aggrecan to aggregate with hyaluronan. In vivo activity of the proteinases was tested by injecting purified HLE and HCG intra-articularly into rabbit stifle joints and quantifying the levels of proteoglycan released into synovial fluids. On a molar basis, HCG was at least tenfold more potent than HLE as a proteoglycanase in vitro. Moreover, HCG was twofold more potent as a proteoglycanase in vivo. In contrast, HLE hydrolyzed elastin approximately 22-fold faster than HCG, but was only slightly more rapid than HCG when [3H]-transferrin was used as substrate. These data indicate that HCG is more potent than HLE as a proteoglycanase both in vitro and in vivo. Thus, HCG could be more important in the pathogenesis of rheumatoid arthritis than previously suspected.

Aggrecans↗
PubMed1993

Adolescent health status measurement: development of the Child Health and Illness Profile.

This report describes the early stages in the development and testing of an instrument, known as the CHIP (Child Health and Illness Profile), for assessing the health of individuals aged 11 through 17. The purpose of the instrument is to assess health in epidemiologic surveys, to determine the existence of systematic differences in health in subpopulations (including the socioeconomically disadvantaged), and to provide a basis for assessing the impact of changes in health services or health policies. An instrument consisting of six domains with 25 subdomains was developed based on the literature, the involvement of focus groups and expert panels, and pretesting in four groups of teenagers known to differ in their health. The results of work with panels of experts suggest that the instrument has content validity. Most domains and subdomains had acceptable reliability as measured by alpha coefficients. Differences in the scores of individuals in the four groups were in the predicted directions, suggesting that the instrument also has construct validity. Additional research is under way to establish other aspects of validity as well as reliability in school populations of adolescents as well as specific clinical settings.

Activities of Daily Living↗
PubMed1992

Inhibition of human serine proteases by substituted 2-azetidinones.

trans-4-Ethoxycarbonyl-3-ethyl-1-(4-nitrophenyl-sulfonyl)-azetidin -3-one described by Firestone et al. (1990, Tetrahedron 46, 2255) as an inhibitor of human leucocyte elastase (HLE) displayed potent, time-dependent inhibition of both HLE and human cathepsin G (Cat-G). The cis-isomer was 7- and 180-fold less active, respectively. The mechanism likely involves opening of the beta-lactam ring by the active site serine to form an acyl-enzyme intermediate(s). This intermediate partitions with ratios of 4:1 between turnover of the inhibitor and formation of relatively stable enzyme-inhibitor complexes from both enzymes. The final HLE-inhibitor complex reactivated with a half-life of 48 h at 25 degrees C and was 16-fold more stable than the Cat-G-inhibitor complex. The stability of the acyl-enzymes supports a "double hit" chemical mechanism involving both serine acylation and alkylation of the histidine. These observations suggest that beta-lactams may be developed as a class of serine protease inhibitors.

Azetidines↗
PubMed1992

Identification of the posttranslational modifications of bovine lens alpha B-crystallins by mass spectrometry.

A combination of mass spectrometric techniques has been used to investigate the amino acid sequence and post-translational modifications of alpha B-crystallin isolated from bovine lenses by gel filtration chromatography and reversed-phase high performance liquid chromatography. Chromatographic fractions were analyzed by electrospray ionization mass spectrometry to determine the homogeneity and molecular weights of proteins in the fractions. The alpha B-crystallin primary gene product, its mono- and diphosphorylated forms, its N- and C-terminal truncated forms, as well as other lens proteins unrelated to the alpha B-crystallins were identified by their molecular weights. Detailed information about the sites of phosphorylation, as well as evidence supporting reassignment of Asn to Asp at position 80, was obtained by analyzing proteolytic digests of these proteins by fast atom bombardment mass spectrometry. Results of this investigation indicate that alpha B-crystallin is phosphorylated in vivo at Ser 45, Ser 59, and either Ser 19 or 21. From the specificity of phosphorylation of alpha-crystallins, it appears that there may be two different kinases responsible for their phosphorylation.

Amino Acid Sequence↗
PubMed1992

Oxidative degradation of phenanthrene by the ligninolytic fungus Phanerochaete chrysosporium.

The ligninolytic fungus Phanerochaete chrysosporium oxidized phenanthrene and phenanthrene-9,10-quinone (PQ) at their C-9 and C-10 positions to give a ring-fission product, 2,2'-diphenic acid (DPA), which was identified in chromatographic and isotope dilution experiments. DPA formation from phenanthrene was somewhat greater in low-nitrogen (ligninolytic) cultures than in high-nitrogen (nonligninolytic) cultures and did not occur in uninoculated cultures. The oxidation of PQ to DPA involved both fungal and abiotic mechanisms, was unaffected by the level of nitrogen added, and was significantly faster than the cleavage of phenanthrene to DPA. Phenanthrene-trans-9,10-dihydrodiol, which was previously shown to be the principal phenanthrene metabolite in nonligninolytic P. chrysosporium cultures, was not formed in the ligninolytic cultures employed here. These results suggest that phenanthrene degradation by ligninolytic P. chrysosporium proceeds in order from phenanthrene----PQ----DPA, involves both ligninolytic and nonligninolytic enzymes, and is not initiated by a classical microsomal cytochrome P-450. The extracellular lignin peroxidases of P. chrysosporium were not able to oxidize phenanthrene in vitro and therefore are also unlikely to catalyze the first step of phenanthrene degradation in vivo. Both phenanthrene and PQ were mineralized to similar extents by the fungus, which supports the intermediacy of PQ in phenanthrene degradation, but both compounds were mineralized significantly less than the structurally related lignin peroxidase substrate pyrene was.

Basidiomycota↗
PubMed1992

Implications on laboratory workload of breast cancer screening.

AIMS: To determine whether the recommended methods for the handling of breast excision biopsy specimens from screening patients cause more laboratory work than those used for non-screening patients. METHODS: All breast excision biopsy specimens from 1990 were identified. Ninety one came from patients identified during the prevalent round of breast screening. These were compared with 127 from non-screening patients operated on by the same surgeon. The workload in terms of initial blocks taken, cases which needed extra blocks or other additional work, and the time taken for delivery of slides to the pathologist were assessed. RESULTS: The screening cases required significantly more initial blocks than the non-screening cases (8.03 v 4.95; p = 0.000001). When looking at the malignant diagnoses this difference was maintained (7.74 v 6.02; p = 0.00014). CONCLUSIONS: Excision biopsy specimens from screening patients require more laboratory work per case than those from non-screening patients. The reasons for these differences lie in the nature of the specimens and their subsequent diagnoses, and in the initial indications for biopsy.

Biopsy↗
PubMed1992

Daily living programme.

Explore the source record for details and available documents.

Community Mental Health Services↗
PubMed1991

Ring fission of anthracene by a eukaryote.

Ligninolytic fungi are unique among eukaryotes in their ability to degrade polycyclic aromatic hydrocarbons (PAHs), but the mechanism for this process is unknown. Although certain PAHs are oxidized in vitro by the fungal lignin peroxidases (LiPs) that catalyze ligninolysis, it has never been shown that LiPs initiate PAH degradation in vivo. To address these problems, the metabolism of anthracene (AC) and its in vitro oxidation product, 9,10-anthraquinone (AQ), was examined by chromatographic and isotope dilution techniques in Phanerochaete chrysosporium. The fungal oxidation of AC to AQ was rapid, and both AC and AQ were significantly mineralized. Both compounds were cleaved by the fungus to give the same ring-fission metabolite, phthalic acid, and phthalate production from AQ was shown to occur only under ligninolytic culture conditions. These results show that the major pathway for AC degradation in Phanerochaete proceeds AC----AQ----phthalate + CO2 and that it is probably mediated by LiPs and other enzymes of ligninolytic metabolism.

Anthracenes↗
PubMed1991

Lipoylation of the E2 components of the 2-oxo acid dehydrogenase multienzyme complexes of Escherichia coli.

The number of functional lipoyl groups in the dihydrolipoyl acetyltransferase (E2) chain of the pyruvate dehydrogenase multienzyme complex from Escherichia coli has been re-assessed by means of a combination of protein-chemical and mass-spectrometric techniques. (1) After the complex had been treated with N-ethyl[2,3-14C]maleimide in the presence of pyruvate, the lipoyl domains were excised from the complex, treated with NaBH4 and re-exposed to N-ethyl[2,3-14C]maleimide. All the chemically reactive lipoyl groups in the native complex were found to be catalytically active. (2) Proteolytic digests of the separated lipoyl domains were examined for the presence of the lipoylation-site peptide, GDKASME, with and without the lipoyl group in N6-linkage to the lysine residue. Only the lipoylated form of the peptide was detected, suggesting that all three lipoyl domains are fully substituted at this site. (3) The behaviour of each lipoyl domain was examined on ion-exchange chromatography in response to alkylation with 4-vinylpyridine after either chemical reduction of the lipoyl group with dithiothreitol or reductive acetylation by the pyruvate dehydrogenase complex in the presence of pyruvate. All three domains exhibited a quantitative shift in retention time, confirming that each domain was fully substituted by an enzymically reactive lipoyl group. (4) When subjected to electrospray mass spectrometry, each domain gave a mass consistent with a fully lipoylated domain, and no aberrant substitution of the target lysine residue was detected. The same result was obtained for the lipoyl domain from the E. coli 2-oxoglutarate dehydrogenase complex. (5) Previous widespread attempts to assess the number of functional lipoyl groups in the pyruvate dehydrogenase multienzyme complex, which have led to the view that a maximum of two lipoyl groups per E2 chain may be involved in the catalytic mechanism, are in error.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
PubMed1991

Elucidation of the primary structures of proteins by mass spectrometry.

A combination of new mass spectrometric methods that can be used to determine the primary structures of proteins, including post-translational modifications, with unprecedented speed and accuracy is described. Structural characterization of alpha-crystallins from bovine lenses has been used to illustrate the methods. The molecular weights of alpha-crystallins fractionated, but not to homogeneity, by reversed-phase HPLC were determined with an uncertainty of 0.01% which is at least 100 times more accurate than is possible using conventional methods. This information was used to identify the primary gene product as well as its phosphorylated and truncated forms. Molecular weight maps of proteolytic digests of these proteins were determined by directly coupled capillary HPLC fast atom bombardment-mass spectrometry. From these maps, the entire amino acid sequence was confirmed, and the phosphorylated peptide was identified. The MS/MS daughter ion mass spectrum of the phosphorylated peptide provided sufficient information to determine which residue was phosphorylated. Because protein structure, including post-translational modifications, is determined on the basis of molecular weight, this method has broad application and will be useful for a variety of diverse and challenging problems in protein structure elucidation.

Amino Acid Sequence↗
PubMed1991

An acyl-carrier-protein-thioesterase domain from the 6-deoxyerythronolide B synthase of Saccharopolyspora erythraea. High-level production, purification and characterisation in Escherichia coli.

The C-terminal region of a multifunctional polypeptide from the 6-deoxyerythronolide B synthase of Saccharopolyspora erythraea is predicted to contain an acyl carrier protein and a thioesterase or acyltransferase activity [Cortes, J., Haydock, S. F., Roberts, G. A., Bevitt, D. J. & Leadlay, P. F. (1990) Nature 348, 176-178]. Site-directed mutagenesis by means of the polymerase chain reaction was used to construct an efficient pT7-based expression plasmid for this domain. The recently developed technique of electrospray mass spectrometry was used to demonstrate that the purified protein had not been post-translationally modified by attachment of a 4'-phosphopantetheine group. However, treatment with the serine proteinase inhibitor phenylmethylsulphonyl fluoride led to highly selective labelling of the predicted active site of the thioesterase or acyltransferase.

Amino Acid Sequence↗
PubMed1991

Transvenous occlusion of clinical and subclinical varicoceles.

Forty-four subfertile men with a palpable or clinically suspected varicocele underwent physical examination, nuclear scan, and testicular venography. The ability of these tests to detect a varicocele was compared. The sensitivity of physical examination was 0.94 on the left and 0.35 on the right. Specificity was 0.22 on the left and 0.77 on the right. The sensitivity of nuclear scan was 0.97 on the left and 0.73 on the right. Specificity was 0.56 on the left and 0.50 on the right. Patients with varicoceles demonstrated by venography were treated with testicular vein embolization. There were no complications except two asymptomatic subintimal injections, both of which occurred on the right side. A total of 19 patients had pre- and post-embolization semen analyses performed. The total motile sperm count increased from 27.0 million to 35.5 million. The overall pregnancy rate in the embolization-treated group was 43.7 percent.

Adult↗
PubMed1991

Milk composition in an American marsupial (Monodelphis domestica).

1. Twenty one sequential milk samples from an American marsupial (Monodelphis domestica) were analyzed. 2. The solids fraction was relatively low initially (10% w/w) and then increased to about 30% at mid-lactation where it remained until lactation ceased. 3. During the first 50 days of lactation the relative proportions of protein, fat and carbohydrate were 36, 28 and 34%, respectively, of the solids fraction. 4. Sodium and magnesium concentrations were constant; however levels of potassium and magnesium varied substantially. 5. The milk of M. domestica exhibits a similar pattern of quantitative and qualitative changes to that seen in Australian marsupials.

Animals↗