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J R Barr

Publications and source records attributed to J R Barr.

16 recordsLinked to original sources

Structural classification of carbohydrates in glycoproteins by mass spectrometry and high-performance anion-exchange chromatography.

A general strategy has been developed for determining the structural class (oligomannose, hybrid, complex), branching types (biantennary, triantennary, etc.), and molecular microheterogeneity of N-linked oligosaccharides at specific attachment sites in glycoproteins. This methodology combines mass spectrometry and high-performance anion-exchange chromatography with pulsed amperometric detection to take advantage of their high sensitivity and the capability for analysis of complex mixtures of oligosaccharides. Glycopeptides are identified and isolated by comparative HPLC mapping of proteolytic digests of the protein prior to, and after, enzymatic release of carbohydrates. Oligosaccharides are enzymatically released from each isolated glycopeptide, and the attachment site peptide is identified by fast atom bombardment mass spectrometry (FAB-MS) of the mixture. Part of each reaction mixture is then permethylated and analyzed by FAB-MS to identify the composition and molecular heterogeneity of the carbohydrate moiety. Fragment ions in the FAB mass spectra are useful for detecting specific structural features such as polylactosamine units and bisecting N-acetylhexosamine residues, and for locating inner-core deoxyhexose residues. Methylation analysis of these fractions provides the linkages of monomers. Based on the FAB-MS and methylation analysis data, the structural classes of carbohydrates at each attachment site can be proposed. The remaining portions of released carbohydrates from specific attachment sites are preoperatively fractionated by high-performance anion-exchange chromatography, permethylated, and analyzed by FAB-MS. These analyses yield the charge state and composition of each peak in the chromatographic map, and provide semiquantitative information regarding the relative amounts of each molecular species. Analytically useful data may be obtained with as little as 10 pmol of derivatized carbohydrate, and fmol sensitivity has been achieved. The combined carbohydrate mapping and structural fingerprinting procedures are illustrated for a recombinant form of the CD4 receptor glycoprotein.

Amino Acid Sequence

Protein and carbohydrate structural analysis of a recombinant soluble CD4 receptor by mass spectrometry.

The primary structure of a soluble form of the CD4 receptor (sCD4) expressed in Chinese hamster ovary cells has been confirmed by mass spectrometric peptide mapping and and tandem mass spectrometry. These studies corroborated 95% of the 369-amino acid-long sequence and established the fidelity of translation of the NH2 and COOH terminal including the absence of "ragged ends." The arrangement of the three disulfide bonds in recombinant sCD4 was also established by mass spectrometry and comparative high performance liquid chromatography mapping and shown to be identical to that expected from previous studies of intrachain disulfide bonding in T4 antigens derived from sheep and mouse. No other arrangements of disulfides were detected. Carbohydrate mapping by mass spectrometry was used to establish that both potential Asn-linked glycosylation sites in sCD4 (Asn271 and Asn300) have oligosaccharides attached. Structural characterization by mass spectrometry and methylation analysis of the heterogeneous family of oligosaccharides at each of the specific attachment sites indicates that the major components of both families of oligosaccharides have the following biantennary structures: (Formula, see text) where m + n = 0-2, and x = 0,1. Minor carbohydrate components having three N-acetylneuraminic acid (NeuAc) groups and an additional hexose-hexosamine unit were detected by high performance anion-exchange chromatography.

Amino Acid Sequence

Clear-cell epithelial neoplasms of the large intestine.

We report four cases of primary clear-cell adenoma and adenocarcinoma of the large intestine. The neoplasms grossly resembled ordinary colonic adenomas and adenocarcinomas but microscopically were composed of uniform cells with optically clear cytoplasm. Mucin stains were negative, and the clear nature of the cytoplasm was due to glycogen accumulation. Areas of transition between normal colonic epithelial constituents and the clear-cell lesion were observed. Three of the four cases stained strongly positively for carcinoembryonic antigen. These lesions are apt to give rise to considerable diagnostic confusion and, in particular, resemble metastatic renal cell carcinoma. The usual strong positive carcinoembryonic antigen reaction is helpful in establishing this diagnosis.

Acid Phosphatase

Classification of human tumours by high-resolution magnetic resonance spectroscopy.

Malignant tumours yield a high-resolution proton magnetic resonance (MR) spectrum. Fifty-one tumour biopsy specimens from patients with cancer of the ovary or colon were examined by magnetic resonance spectroscopy (MRS) to determine whether it was possible to identify tumour subtypes with metastatic potential. Relaxation parameters (T2) for visible lipid methylene protons were within the range of those measured for three animal metastasis models. Primary carcinomas with metastases gave T2 values greater than 350 ms, whereas carcinomas not associated with known metastases at the time of tumour excision gave a range of values from 150 to 1500 ms. All but two carcinomas gave a long T2 (greater than 350 ms) indicating metastatic potential. The MRS method designated five of six histologically borderline epithelial ovarian tumours as malignant with metastatic potential. MRS may be sensitive enough to detect malignant cells in a tumour which is of intermediate or borderline malignancy by light microscopy. Malignancy without a potential for metastasis is uncommon.

Colonic Neoplasms

The clinical recognition of cystic carcinoma of the breast.

Cystic malignant growths of the breast are uncommon, being only 1 per cent of all carcinomas of the breast. They can be easily diagnosed clinically by noting an unusual patient age, the nature of the fluid aspirated (bloody or peculiar) and recurrence or persistence of the "cyst." The results of cytologic examination of the aspirate has not been helpful in the diagnosis of these lesions.

Age Factors

Cystadenoma and cystadenocarcinoma of the pancreas.

Cystic neoplasms of the pancreas are uncommon. The authors present one case each of a cystic carcinoma (possibly acinar, which is a rare variant of pancreatic carcinoma), cystadenocarcinoma arising in a mucinous cystadenoma and cystadenoma. These cases illustrate important features of the natural history and management of these lesions. The macrocystic cystadenoma is generally regarded as premalignant and merits aggressive treatment, while the microcystic type is considered benign. If the cystadenoma is removed completely, there should be no recurrence. The cystadenocarcinoma differs from duct carcinoma; usually, the tumour can be removed, and when this is possible the prognosis is excellent. Both the cystadenoma and the cystadenocarcinoma can be mistaken for a pancreatic pseudocyst. The distinction is important.

Adult

Angiosarcoma of serous membranes.

Three angiosarcomas of serous membrane are described. One originated in peritoneum, one in pleura, and one in pericardium. The tumor arising in peritoneum was a cystic lymphangiosarcoma and may have been caused by previous therapeutic irradiation for carcinoma of the endometrium. This tumor invaded the peritoneal and right pleural cavities extensively and thus resembled diffuse mesothelioma in its behavior. The tumors arising in pleura and pericardium were hemangiosarcomas.

Aged

Strongyloides stercoralis.

Strongyloides stercoralis, a nematode not well known to many Canadian physicians, infects 35% of some tropical populations. Larvae can be isolated from the stools in 25% of cases and from duodenal aspirates in 95%. Treatment is with thiabendazole given twice daily in a dose of 25 mg/kg up to a maximum of 1.5 g/d. Frenquently an individual with a previously asymptomatic infection presents with hyperinfection and death rapidly ensues, but usually classical symptoms are present. Such a case is described. Immunosuppression is frequently associated with the hyperinfective state.

Canada

Elastofibroma.

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Aged

5-Alkylresorcinols from Hakea amplexicaulis that cleave DNA.

A dichloromethane extract of Hakea amplexicaulis was found to cause strand scission of phi X174 replicative form DNA in the presence of Cu(II). Bioassay-guided fractionation of this extract afforded five compounds capable of mediating DNA relaxation. Structure determination of the active principles indicated that they were 5-tridecylresorcinol (1), 5-pentadec-cis-8-enylresorcinol (2), 5-heptadeca-8,11-dienylresorcinol (3), 5-pentadecylresorcinol (4), and 5-heptadec-cis-8-enylresorcinol (5). As noted previously for compounds in this structural series, DNA cleavage was enhanced significantly by incubation under (alkaline) conditions known to promote oxygenation of the parent compounds on the aromatic nucleus.

Alkylation