Structure-function relationship in glycosylated alpha-chymotrypsin as probed by IMAC and IMACE.
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Chemical glycosylation of bovine alpha-chymotrypsin, by a glucosamine adduct on the carboxyl group, results in the modification of its catalytic activity. The structural alterations of alpha-chymotrypsin resulting from its glycosylation are studied by immobilized metal-ion affinity chromatography (IMAC) and immobilized metal-ion affinity capillary electrophoresis (IMACE). The chemical glycosylation of alpha-chymotrypsin generates two distinct subpopulations of the protein: one which totally loses the initial affinity for IDA-Cu(II) and another which exhibits an increased affinity for the metal chelate ligand. (+info)
OBJECTIVE: Visualization of enterogastric reflux (EGR) may be present during hepatobiliary imaging. Reflux of bile may damage the gastric mucosa, altering its function, and cause such symptoms as epigastric pain, heartburn, nausea, intermittent vomiting and abdominal fullness. These symptoms also are associated with gallbladder disease. The aim of this study was to quantitate the EGR index (EGRI) and to determine if a difference exists in normal and abnormal responses using standard cholecystokinin (CCK)-augmented hepatobiliary imaging. METHODS: This study used 129 patients. LAO dynamic data on a 128 x 128 matrix at a rate of 1 frame/min were obtained. After the gallbladder ejection fraction (GBEF) was determined, the EGRI (%) was calculated by relating the counts in the gastric ROI to the counts in the hepatobiliary ROI at a specified time. The results were compared with the patient's final clinical diagnosis. RESULTS: Normal responders (GBEF > or = 35%) had a higher EGRI than abnormal responders with a P = 0.001 EGR observed in 75 patients (58.1%). Significant reflux (EGRI > or = 14.2% at 15 min) was observed in 29 additional patients (22.5%). Patients with EGRI > or = 24.5% showed a strong association with the pathophysiologic syndrome of gastritis, alkaline reflux, gastric ulcer and gastro esophageal reflux disease. There was no EGR observed in the remaining 25 patients (19.4%). CONCLUSION: This simple addition to the CCK-augmented hepatobiliary imaging may both detect and quantitate abnormal EGR as the cause of the patient's symptoms in the presence of a normal GBEF result, and/or those patients with risk factors for gastritis. (+info)
The role of morphine-augmented cholescintigraphy and real-time ultrasound in detecting gallbladder disease.
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OBJECTIVE: Rapid diagnosis of acute cholecystitis is essential to minimize morbidity and mortality. The purpose of this study was to assess the diagnostic utility of cholescintigraphy using morphine augmentation compared with ultrasound, in acute and chronic gallbladder disease. METHODS: Cholescintigrams were performed on 103 patients suspected of having acute cholecystitis. In 79 patients (Group A) morphine sulfate was administered to reduce the scintigraphic imaging time if the gallbladder was not visualized during the first hour. In 24 control patients (Group B) no morphine was administered. All patients were evaluated clinically and 93 patients had concurrent ultrasound examination. RESULTS: The clinical presentation was nonspecific. The ultrasound findings were sensitive in detecting gallbladder disease (100%), but had low specificity (24%). Only findings of sediments and pericholecystic fluid were specific for cystic duct obstruction. Morphine augmentation reduced the imaging time by 126 min in patients with chronic cholecystitis. CONCLUSION: Real-time ultrasound has low specificity for gallbladder disease. In the presence of an abnormal ultrasound, it is essential to perform a hepatobiliary scan, either to exclude gallbladder disease or distinguish acute from chronic cholecystitis. Low-dose morphine administration is a safe and useful adjunct to standard cholescintigraphy by substantially reducing the time required to obtain a diagnostic study. (+info)
Efficacy of morphine sulfate-augmented hepatobiliary imaging in acute cholecystitis.
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OBJECTIVE: A review of the English language literature was performed to determine the sensitivity and specificity of morphine sulfate-augmented hepatobiliary imaging for acute cholecystitis. Twenty publications, involving 914 patients, were reviewed from journals published between 1984 and 1999. The analysis of these patients has resulted in the largest combined review study to date. The sensitivity and specificity of morphine-augmented hepatobiliary imaging were calculated to be 96.1% and 88.6%, respectively. After reading this paper, the nuclear medicine technologist should be able to: (a) discuss the clinical use of morphine augmentation during hepatobiliary imaging; and (b) state the sensitivity and specificity of morphine sulfate-augmented hepatobiliary imaging. (+info)
Identification, purification, and characterization of iminodiacetate oxidase from the EDTA-degrading bacterium BNC1.
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Microbial degradation of synthetic chelating agents, such as EDTA and nitrilotriacetate (NTA), may help immobilizing radionuclides and heavy metals in the environment. The EDTA- and NTA-degrading bacterium BNC1 uses EDTA monooxygenase to oxidize NTA to iminodiacetate (IDA) and EDTA to ethylenediaminediacetate (EDDA). IDA- and EDDA-degrading enzymes have not been purified and characterized to date. In this report, an IDA oxidase was purified to apparent homogeneity from strain BNC1 by using a combination of eight purification steps. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single protein band of 40 kDa, and by using size exclusion chromatography, we estimated the native enzyme to be a homodimer. Flavin adenine dinucleotide was determined as its prosthetic group. The purified enzyme oxidized IDA to glycine and glyoxylate with the consumption of O2. The temperature and pH optima for IDA oxidation were 35 degrees C and 8, respectively. The apparent Km for IDA was 4.0 mM with a kcat of 5.3 s(-1). When the N-terminal amino acid sequence was determined, it matched exactly with that encoded by a previously sequenced hypothetical oxidase gene of BNC1. The gene was expressed in Escherichia coli, and the gene product as a C-terminal fusion with a His tag was purified by a one-step nickel affinity chromatography. The purified fusion protein had essentially the same enzymatic activity and properties as the native IDA oxidase. IDA oxidase also oxidized EDDA to ethylenediamine and glyoxylate. Thus, IDA oxidase is likely the second enzyme in both NTA and EDTA degradation pathways in strain BNC1. (+info)
A new crosslinker for mass spectrometric analysis of the quaternary structure of protein complexes.
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Mass spectrometric structural analysis of crosslinked peptides is a powerful method to elucidate the spatial arrangement of polypeptides in protein complexes. Our aim is to develop bifunctional crosslinkers that, after crosslinking protein complexes followed by proteolytic digestion, give rise to crosslinked peptides that can be readily tracked down by mass spectrometry. To this end we synthesized the crosslinker N-benzyliminodiacetoyloxysuccinimid (BID), which yields stable benzyl cation marker ions upon low-energy collision-induced dissociation (CID) tandem mass spectrometry. Sensitive detection of the marker ion upon low-energy CID is demonstrated with different BID-crosslinked peptide preparations. With BID it becomes possible to retrieve crosslinked and crosslinker-adducted peptides, without the necessity of purifying crosslinked peptides prior to identification. The basic design of this crosslinker can be varied upon, in order to meet specific crosslinking needs. (+info)
Polymer properties on resins composed of UDMA and methacrylates with the carboxyl group.
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The properties of dental matrix resins have been improved by synthesis of new monomers. However, except for improvements in water-resistance, monomers with better mechanical properties than Bis-GMA and UDMA could not being synthesized. Changing the point of emphasis, we tried to improve the mechanical properties controlling the matrix resin higher structure using noncovalent bonds. We prepared a matrix resin structured by UDMA, which is a high viscosity base monomer with imino groups, and by a low viscosity acidic monomer with carboxyl groups, which permits noncovalent bonds such as hydrogen bonds or electrostatic interaction with imino groups. The maximal mechanical strength for matrix resins structured by UDMA and an acidic monomer was obtained with a composition of imino groups and carboxyl groups at a ratio of 1:1. This mechanical strength value was higher than those obtained with UDMA resin or with a Bis-GMA/TEGDMA/UDMA resin with typical composition. The improvement in mechanical properties may be due to the complex based on noncovalent bonds, between the imino groups of UDMA and the carboxyl groups of the acidic monomers. (+info)
99mTc-mebrofenin scintigraphy for evaluating liver disease in a rat model of Wilson's disease.
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The purpose of this study was to establish whether (99m)Tc-mebrofenin could noninvasively assess liver function in Wilson's disease. METHODS: Long-Evans Cinnamon (LEC) rats, which reproduce Wilson's disease with copper toxicosis, and their normal counterparts, Long-Evans Agouti (LEA) rats, were studied. Scintigraphic findings were correlated with biliary mebrofenin excretion and residual organ counts and with hepatic copper content, histology, copper excretion capacity, and liver test results. RESULTS: Serum alanine aminotransferase (ALT) levels were elevated in some LEC rats, whereas serum bilirubin levels were normal. Liver histology was normal in LEA rats, whereas LEC rats showed multiple abnormalities. Mebrofenin was incorporated rapidly in LEA rats, with a mean time to peak liver activity of 80 +/- 30 s, followed by prompt biliary excretion of the tracer. In LEC rats, the mean time to peak activity, 283 +/- 190 s, was significantly longer (P = 0.001). The time to half of peak activity, indicating tracer clearance, was significantly greater in LEC rats than in LEA rats (1,825 +/- 1,642 s vs. 524 +/- 82 s, P = 0.002). Hepatic mebrofenin handling correlated with hepatic copper content, histologic grade, copper excretion capacity, and serum ALT. CONCLUSION: Correlation of (99m)Tc-mebrofenin handling with liver morphology, function, and copper accumulation in LEC rats suggests that mebrofenin scintigraphy can be useful for noninvasively monitoring disease progression and therapeutic response in Wilson's disease. Although the data were obtained in an animal model of Wilson' disease, these biochemical parameters likely reflect liver damage in general, suggesting that there may be a role for mebrofenin scintigraphy in other chronic liver diseases as well. (+info)