Immunochemical specificity of antisera raised against the synthetic encephalitogenic peptide SH624, residues 59–74 of the myelin basic protein

Synthetic peptide SH624 (SHHPARTAHYGSLPQK), residues 59–74 of human myelin basic protein (MBP) was found to be encephalitogenic in the rabbit. Four antisera raised, against the peptide were employed in a liquid-phase equilibrium competitive radioimmunoassay with a series of synthetic peptide analogs of the region to probe the structural requirements of the B-cell determinant subsumed within SH624. The cross-reactivities of the four antisera with intact MBP were also examined. Immunochemical analyses of the four antisera suggested specificities directed against a conformational determinant dependent upon residues from the more phylogenetically conserved carboxyl C-terminal region, residues 65–74 (TAHYGSLPQK) of the synthetic immunogen. Peptide analogs shorter than SH624 from the C-terminal end showed no cross-reactivity with any of the reagent antisera while analogs shorter from the N-terminal end and including the encephalitogenic sequence TTHYGSLPQK, as well as, HYGSLPQK were reactive under equilibrium competitive conditions. SH624-reactive antibodies, cross-reactive with purified heterologous MBPs from 10 different species were also identified in all four reagent antisera. The results of these experiments support previous investigations demonstrating the accessibility of the encephalitogenic 65–74 region in intact MBP. They also underscore the importance of B-cell recognition of organ specific antigenic determinants with respect to MBP immunology and, in particular, the recognition of autoreactive determinants in the neighborhood of encephalitogenic centers.     

Homozygous paracentric inversion 12 in a mentally retarded boy: a case report and review of the literature

Summary A mentally retarded male was found to be homozygous for a paracentric inversion of the long arm of chromosome 12(inv(12)(q21.1q23.2)). His parents, who are first cousins, and his phenotypically normal younger brother are inversion heterozygotes. Homozygous structural rearrangements are discussed and cases of paracentric inversions, including a further nine previously unpublished, are reviewed.     

A natural hybrid newt, Triturus helveticus×T. vulgaris, from a pond in mid-Wales

The first unequivocal example of a natural Trilurus helvelicus × T. vulgaris hybrid is described. The specimen was a male and discriminant analysis of physical characters indicated that it was morphologically intermediate between the parent species. A karyotype confirmed that the hybrid bore a haploid set of chromosomes from T. helveticus and a haploid set from T. vulgaris. Examination of the sex chromosomes showed that it was the result of mating between a male T. helveticus and a female T. vulgaris. As numerous mature sperm bundles were observed in both testes, the hybrid was therefore potentially fertile.     

The response of murine intestinal crypts to short-range promethium-147 beta irradiation: deductions concerning clonogenic cell numbers and positions

An exteriorized loop of mouse intestine was exposed to 147Pm low-energy electrons, where the dose rate decreased by a factor of 5 from the base of the crypt to the top of the proliferative zone. A crypt survival curve was obtained, expressed in terms of exposure time. The shape of the curve was interpreted in terms of survival parameters for colony-forming cells (clonogens) derived using 137Cs gamma rays and the depth-dose curve measured for 147Pm electrons. It is concluded that the shape of the crypt survival curve using 147Pm electrons is inconsistent with the notion of either the presence of a large number of clonogens or a small number near the top of the proliferative zone. A computer fitting procedure showed that the best agreement between predicted and observed curves was achieved with 2.7 +/- 0.5 clonogens at cell position 5.6 +/- 0.6, in the putative stem-cell zone.     

Approximate matching of regular expressions

Given a sequenceA and regular expressionR, theapproximate regular expression matching problem is to find a sequence matchingR whose optimal alignment withA is the highest scoring of all such sequences. This paper develops an algorithm to solve the problem in timeO(MN), whereM andN are the lengths ofA andR. Thus, the time requirement is asymptotically no worse than for the simpler problem of aligning two fixed sequences. Our method is superior to an earlier algorithm by Wagner and Seiferas in several ways. First, it treats real-valued costs, in addition to integer costs, with no loss of asymptotic efficiency. Second, it requires onlyO(N) space to deliver just the score of the best alignment. Finally, its structure permits implementation techniques that make it extremely fast in practice. We extend the method to accommodate gap penalties, as required for typical applications in molecular biology, and further refine it to search for substrings ofA that strongly align with a sequence inR, as required for typical data base searches. We also show how to deliver an optimal alignment betweenA andR in onlyO(N+logM) space usingO(MN logM) time. Finally, anO(MN(M+N)+N ²logN) time algorithm is presented for alignment scoring schemes where the cost of a gap is an arbitrary increasing function of its length.     

A redetermination of the concentration of alpha 2-macroglobulin in human plasma

The concentration of alpha 2-macroglobulin in human plasma has been remeasured utilizing a carefully isolated and characterized sample of alpha 2-macroglobulin as a standard. A highly purified sample of alpha 2-macroglobulin with a total trypsin binding capacity of 1.7 mol trypsin/mol alpha 2-macroglobulin was used as a standard for both a radial immunodiffusion and a rocket immunoelectrophoresis technique. With this preparation as a standard, the concentration of alpha 2-macroglobulin in a normal plasma pool over 10,000 donors was found to be about 1.2 mg/ml. A similar concentration (1.3 mg/ml) was found when using a functional trypsin binding assay. This concentration is considerably less than the usually accepted mean of the normal range for alpha 2-macroglobulin.     

Degradation of human proteoglycan aggregate induced by hydrogen peroxide. Protein fragmentation, amino acid modification and hyaluronic acid cleavage.

We have previously shown that treatment of neonatal human articular-cartilage proteoglycan aggregates with H2O2 results in loss of the ability of the proteoglycan subunits to interact with hyaluronic acid and in fragmentation of the link proteins [Roberts, Mort & Roughley (1987) Biochem. J. 247, 349-357]. We now show the following. (1) Hyaluronic acid in proteoglycan aggregates is also fragmented by treatment with H2O2. (2) Although H2O2 treatment results in loss of the ability of the proteoglycan subunits to interact with hyaluronic acid, the loss of this function is not attributable to substantial cleavage of the hyaluronic acid-binding region of the proteoglycan subunits. (3) In contrast, link proteins retain the ability to bind to hyaluronic acid following treatment with H2O2. (4) The interaction between the proteoglycan subunit and link protein is, however, abolished. (5) N-Terminal sequence analysis of the first eight residues of the major product of link protein resulting from H2O2 treatment revealed that cleavage occurred between residues 13 and 14, so that the new N-terminal amino acid is alanine. (6) In addition, a histidine (residue 16) is converted into alanine and an asparagine (residue 21) is converted into aspartate by the action of H2O2. (7) Rat link protein showed no cleavage or modifications in similar positions under identical conditions. (8) This species variation may be related to the different availability of histidine residues required for the co-ordination of the transition metal ion involved in hydroxyl-radical generation from H2O2. (9) Changes in function of these structural macromolecules as a result of the action of H2O2 may be consequences of both fragmentation and chemical modification.     

Phosphate promotes glycation of antithrombin III which interferes with heparin binding

Nonenzymatic glycation of antithrombin III has been reported to cause the reduction of heparin-catalyzed thrombin-inhibiting activity in diabetes. The effect of in vitro nonenzymatic glycation of pure antithrombin III on heparin binding and heparin-potentiated activity under a variety of buffers and pH values was studied to further clarify the physiological significance of this reaction. The extent of glycation, measured by the fructosamine assay and [14C]glucose binding, was enhanced by the presence of phosphate ion (pH 7.45, 8.5 and 9.5) and increased linearly with increasing phosphate ion concentration from 0.01 to 0.2 M phosphate. Conversely, the heparin-catalyzed antithrombin activity decreased from 93.1% of controls for 0.01 M phosphate to 73.5% for 0.2 M phosphate as the extent of glycation increased. The increase in intrinsic fluorescence induced by binding of heparin to antithrombin III was also moderated by glycation of antithrombin III in a dose-dependent manner with a negative correlation coefficient of -0.94. Direct measurement of the heparin binding by affinity chromatography showed a decrease in the heparin-binding fraction which correlated with the degree of glycation and the decrease in heparin-catalyzed activity. These studies suggest that nonenzymatic glycation may be responsible for the reduction in antithrombin III activity observed in some diabetics.     

Detection of prophospholipase A2 in human spermatozoa

Prophospholipase A2 (proPA2) has been isolated from human spermatozoa after acid extraction and chromatography on hydrophobic WP-Butyl (C4) and ion-exchange (SP 5PW) columns. The addition of benzamidine, a noncompetitive synthetic trypsin inhibitor, to semen samples has kept a portion of the sperm phospholipase A2 (PA2) in its zymogen form and allowed its isolation after acid extraction. When radioactive phosphatidylcholine (PC) or phosphatidylethanolamine (PE) were used as substrates, an identical elution profile of this enzyme was obtained on a C4 column. The proenzyme was separated from active PA2 on the C4 column. Human sperm proPA2 exhibited a less cationic charge than active PA2 on the SP 5PW column. Porcine pancreatic proPA2 had the same chromatographic behavior on high performance liquid chromatography (HPLC) (SP 5PW) as human sperm proPA2. The purification procedure resulted in the isolation of proPA2 which, upon activation by proteolysis, presented the same chromatographic elution profile on HPLC as active PA2 of human spermatozoa and porcine pancreas. Thus, a zymogen form of PA2 exists in human spermatozoa.     

Use of the doubly labeled water method for measurement of energy expenditure, total body water, water intake, and metabolizable energy intake in humans and small animals

The basis of the doubly labeled water method is measurement of the differential rates of disappearance of two isotopes of water (H2 18O and either 2H2O or 3H2O, administered at the start of the study) from body water. Published studies indicate that, in its current forms, this technique can be used to provide accurate and reasonably precise information on carbon dioxide production, total body water, and water intake in free-living humans and many small animals. Total energy expenditure can be calculated from carbon dioxide production with little loss of precision. Metabolizable energy intake can also be predicted, as the sum of total energy expenditure plus an estimate for the change in body energy stores during the measurement, but this prediction is unlikely to be accurate and precise unless the subject is in approximate energy balance.