A novel method of multiple alignment of biopolymer sequences

A novel algorithm for multiple alignment of biological sequences is suggested. At the first step the DotHelix procedure is employed for construction of motifs, i.e. continuous fragments of local similarity of various “thickness” and strength, and then these motifs are concatenated into chains consistent with the order of letters in the sequences. The algorithm is implemented in the MA-Tools program of the GeneBee package. An example illustrating the effectivity of the algorithm is presented.     

Identification and characterization of two novel lymphocyte function-associated antigens, L24 and L25

We describe the function and cell distribution of two novel cell surface antigens, L24 and L25. These antigens are broadly distributed on human lymphocytes. Monoclonal antibodies specific for these molecules block lysis by Class I- and II-specific cytotoxic T lymphocytes, but do not affect any other T cell functions tested. Anti-L24 antibody immunoprecipitates a molecule composed of two disulfide-linked monomers of 140 kd each. Anti-L25 antibody immunoprecipitates three proteins of 150, 85, and 75 kd. The study of these and other function associated molecules may provide insight into mechanisms of cytotoxic T lymphocyte recognition and/or function.     

In vivo environmental temperature and the in vitro pattern of luminal acidification in turtle bladders. Evidence for HCO3 ion reabsorption

In this study, it is shown how to transfer tared aliquots of (HCO3 + CO2)-containing luminal fluids directly into the mercury-sealed chamber of a modified Van Slyke apparatus and how to obtain direct as well as indirect manometric determinations of dissolved CO2 ([CO2]f) in each aliquot of such fluids. It is next shown that the pattern of in vitro luminal acidification in an isolated turtle bladder sac depends upon the prior in vivo ambient temperature to which the donor turtle had become adapted. Under in vivo conditions, the food intake, physical activity, and acid excretion of 32 degrees C-adapted turtles are greater than those of 21 degrees C or 26 degrees C-adapted turtles. Under in vitro conditions of incubating isolated bladder sacs (from 21, 26, and 32 degrees C turtles) in (HCO3 + CO2)-containing Ringer media at a single temperature (21 degrees C), the patterns of luminal acidification are as follows: (a) The rate of depletion of luminal [HCO3] is greatest in bladders from the 32 degrees C-adapted turtles. (b) Concomitant decreases in luminal [CO2]f, [HCO3], and pH (the 'CO2-decreasing patterns' of luminal acidification) develop in all bladders from 32 degrees C turtles, in half of those from 26 degrees C turtles, but in less than one-fifth of those from 21 degrees C-adapted turtles: and (c) a CO2-increasing pattern of luminal acidification is found in most of the bladders from 21 degrees C-adapted turtles. A postulated bicarbonate ion-reabsorbing pump is consistent with all of these patterns of luminal acidification.     

Clathrin structure characterized with monoclonal antibodies. II. Identification of in vivo forms of clathrin

Clathrin was isolated from detergent-solubilized, biosynthetically radiolabeled cells by immunoprecipitation with anti-clathrin monoclonal antibodies. Immunoprecipitates obtained after pulse-chase labeling demonstrated that after biosynthesis the LCa light chain of clathrin could be found either complexed to heavy chain or in a free pool (not associated with heavy chain) which decreased steadily over time. More than half of the free LCa disappeared within the first hour after biosynthesis, but some was still detectable after several hours. Incorporation of clathrin LCa light chain and heavy chain into coated vesicles was coordinate and increased up to 4 h after biosynthesis. Comparison of these kinetics suggested that once incorporated into coated vesicles, LCa and heavy chain did not dissociate, even during depolymerization of the vesicle. There was also little apparent degradation of clathrin found in coated vesicles for up to 22 h after biosynthesis. Immunoprecipitation with anti-clathrin monoclonal antibodies was carried out after fractionation of continuously radiolabeled cell lysates using two different sizing columns. These experiments indicated that the triskelion form of clathrin that has been isolated from coated vesicles in vitro also exists in vivo. They also confirmed the existence of a transient but detectable pool of newly synthesized free LCa light chain.     

Novel peripheral blood-derived human cell lines with properties of megakaryocytes

For 18 mo, we derived 18 cell lines from 11 donors with various clinical profiles ranging from normal to leukemic. Suspension cultures were initiated with 1 X 10(6) mononuclear blood cells/ml of nutrient medium containing 10% human serum and 10% lectin-stimulated human lymphocyte conditioned medium. The cultures were monitored weekly by morphological analyses of Wright-Giemsa-stained cell preparations. All successful cultures showed a significant decline in viability during the first 3-4 wk with rate "lymphoid" cells observed in mitosis. Within the next 2 wk, the proliferating cells gave rise to a rapidly expanding population of mononuclear cells. As the cultures expanded, cell morphology became heterogeneous with respect to cell size and nuclear ploidy, with an accumulation of giant multinuclear cells that were suggestive of megakarocytes. Even though the cells did not have the classical morphology of mature platelet-forming megakaryocytes, 90% of the cells within a cell line were positive by direct or indirect immunofluorescence for the platelet membrane glycoproteins IIb and IIIa; for surface markers HLA-Dr and B2-microglobulin; for intracellular platelet-derived growth factor and platelet factor IV; and for membrane affinity or binding with serum platelet-derived growth factor and platelet factor IV. These results suggest that a blood precursor cell, most likely a primitive megakaryoblast, was isolated from the peripheral blood and was provided with an optimal culture environment for sustained growth. These cells did not mature to a more differentiated stage, perhaps owing to regulatory factor deficiencies in this in vitro system. The remarkable frequency of obtaining cell lines with megakaryocyte properties from normal peripheral blood and the capacity of some normal donors to repeatedly yield these cell lines make this cell culture system indeed unique by being selective for putative megakaryocyte precursors.     

Genome multiplication in cardiomyocytes of fast- and slow-growing mice

The number of myocytes and the percentage of cells with a high degree of ploidy increased in the heart ventricles of fast-growing mice compared with slow-growing ones. The mean incidence of octa- and hexadecaploid (by summary DNA content) myocytes was 7% in the slow-growing and 23% in the fast-growing, weaned mice. In these groups, the total myocyte number varied by 20%. There were 43% more myocyte genomes in the heart ventricles of the fast-growing mice than in those of the slow-growing mice. The same differences in cell number and ploidy persist in 90-day-old mice in spite of feeding ad libitum after weaning.     

Evaluation of the diagnostic application of an enzyme immunoassay for Clostridium perfringens type A enterotoxin

The diagnostic application of an enzyme immunoassay for Clostridium perfringens type A enterotoxin was evaluated. Test results from 100 individuals associated with C. perfringens gastroenteritis outbreaks and 111 control individuals were included. The assay sensitivity was 93.7%, and the assay specificity was 98.7%.