Direct incorporation of glucosamine and N-acetylglucosamine into exopolymers by Gluconacetobacter xylinus (=Acetobacter xylinum) ATCC 10245: production of chitosan-cellulose and chitin-cellulose exopolymers

Gluconacetobacter xylinus (=Acetobacter xylinum) ATCC 10245 incorporated 2-amino-2-deoxy-D-glucose (glucosamine) and 2-acetamido-2-deoxy-D-glucose (N-acetylglucosamine), but not 3-O-methyl-D-glucose or 2-deoxy-D-glucose into exopolymers. Incorporation was confirmed by gas chromatography with and without mass spectrometry, Fourier transform infrared, and 1H nuclear magnetic resonance. The average molar percentage of glucosamine and N-acetylglucosamine in the exopolymers was about 18%.     

A test for linkage and association in general pedigrees: the pedigree disequilibrium test

Family-based tests of linkage disequilibrium typically are based on nuclear-family data including affected individuals and their parents or their unaffected siblings. A limitation of such tests is that they generally are not valid tests of association when data from related nuclear families from larger pedigrees are used. Standard methods require selection of a single nuclear family from any extended pedigrees when testing for linkage disequilibrium. Often data are available for larger pedigrees, and it would be desirable to have a valid test of linkage disequilibrium that can use all potentially informative data. In this study, we present the pedigree disequilibrium test (PDT) for analysis of linkage disequilibrium in general pedigrees. The PDT can use data from related nuclear families from extended pedigrees and is valid even when there is population substructure. Using computer simulations, we demonstrated validity of the test when the asymptotic distribution is used to assess the significance, and examined statistical power. Power simulations demonstrate that, when extended pedigree data are available, substantial gains in power can be attained by use of the PDT rather than existing methods that use only a subset of the data. Furthermore, the PDT remains more powerful even when there is misclassification of unaffected individuals. Our simulations suggest that there may be advantages to using the PDT even if the data consist of independent families without extended family information. Thus, the PDT provides a general test of linkage disequilibrium that can be widely applied to different data structures.     

Absence of significant linkage between phonological coding dyslexia and chromosome 6p23-21.3, as determined by use of quantitative-trait methods: confirmation of qualitative analyses

We recently reported the absence of significant linkage of phonological coding dyslexia (PCD) to chromosome 6p23-p21.3 in 79 families with at least two affected siblings, even though linkage of dyslexia to this region has been found in four other independent studies. Whereas, in our previous analyses, we used a qualitative (affected, unaffected, or uncertain) PCD phenotype, here we report a reanalysis of linkage to the chromosome 6p region, by use of four quantitative measures of reading disability: phonological awareness, phonological coding, spelling, and rapid-automatized-naming (RAN) speed. The phonological-coding and spelling measures were highly correlated with each other and with the qualitative PCD phenotype, whereas the phonological-awareness and RAN-speed measures were only moderately correlated with the other measures. Using two-point and multipoint quantitative-trait sib-pair linkage analyses and variance-components analyses, we were unable to detect significant evidence for a locus in the 6p23-p21.3 region influencing any of the quantitative reading measures, supporting our previous qualitative linkage results. The most likely explanation for our inability to detect linkage between dyslexia and this region is that families with subtypes of dyslexia linked to this region are underrepresented in our sample, because of either chance or varying ascertainment criteria.     

The Replication Initiation Protein Sld2 Regulates Helicase Assembly

Assembly of the Cdc45-Mcm2-7-GINS (CMG) replicative helicase complex must be regulated to ensure that DNA unwinding is coupled with DNA synthesis. Sld2 is required for the initiation of DNA replication in budding yeast. We identified a mutant of Sld2, Sld2-m1,4, that is specifically defective in Mcm2-7 binding. When this sld2-m1,4 mutant is expressed, cells exhibit severe inhibition of DNA replication. Furthermore, the CMG complex assembles prematurely in G₁ in mutant cells, but not wild-type cells. These data suggest that Sld2 binding to Mcm2-7 is essential to block the inappropriate formation of a CMG helicase complex in G₁. We also study a mutant of Sld2 that is defective in binding DNA, sld2-DNA, and find that sld2-DNA cells exhibit no GINS-Mcm2-7 interaction. These data suggest that Sld2 association with DNA is required for CMG assembly in S phase.     

NMR studies of the stable mismatch purine-thymine in the self-complementary d(CGPuAATTTCG) duplex in solution

One- and two-dimensional nuclear Overhauser effect experiments demonstrate that a single hydrogen bond between a T imino proton and purine N3 is sufficient to hold the base pair dPu.dT in d(CGPuAATTTCG) by a Watson-Crick fashion rather than a Hoogsteen type. In addition, the dPu.dT base pair is well stacked with neighboring base pairs. The spin-lattice relaxation measurements at 30 and 35 degrees C of two decamers, d(CGPuAATTTCG) and d(CGAAATTTCG), reveal that the elimination of two single hydrogen bonds of dA.dT base pairs (due to the substitution of adenine for purine) in the sequence results in an increase in the overall imino proton exchange rate from 7 to 36 s-1 at the site of mismatch.     

Effects of X irradiation on angiogenesis.

We have evaluated the effect of X irradiation on the mesenchymal tissue growth (blood capillaries and stromal cells) in an angiogenesis system in the mouse. This was accomplished by implanting a polyvinyl alcohol sponge disc in the subcutis of the thorax, and quantifying the extent of growth reduction of capillaries and stromal cells following graded doses of X rays. The sponge disc contained a centrally located pellet impregnated with 20 micrograms of epidermal growth factor and coated with a thin film of slow-releasing compound. Total growth of vessels and fibroblasts was determined by morphometric analysis of histologic sections. The incorporation of [3H]TdR was measured during a 24-h period. A dose-response relationship was observed when X irradiation was given on Day 11 after implantation, with the disc removed on Day 20. A single dose of 15 Gy reduced both the rate of incorporation of [3H]TdR and the total growth area. These and previous observations point to endothelial cells as important targets of ionizing radiation in the stroma, especially during the period of active proliferation of these cells, induced by growth factors.     

Cytometrically detected specific binding of interleukin 2 to cells.

We have developed a new technique for detecting binding of interleukin 2 (IL-2) to cells. This technique involves incubating the cells with IL-2 and then analysing the cell surface with specific anti-IL-2 antibodies and flow cytometry. This binding was only detected on tumor cells that possessed the p55 subunit of the IL-2 receptor. The role of p55 was ascertained by inhibition of the binding with a monoclonal antibody to p55. Although p55 is necessary for cytometrically detected IL-2 binding, further studies demonstrated that p55 is not sufficient. Thus, cytometrically-detected binding is likely to involved the contribution of other IL-2 surface receptors. Interleukin-2 binding to peripheral blood T lymphocytes and to a non-transformed T-cell clone was also detected cytometrically and it was shown that this binding is regulated by the activation status of the cells. Whereas IL-2 binding to quiescent T cells could not be detected, upon activation abundant binding was seen. The functional consequences of this type of cellular binding were studied. Interleukin-2 binding to cells during a short pulse was shown to have significant long-term consequences both for T-cell proliferation and for the enhancement of major histocompatibility complex (MHC)-non-restricted cytotoxicity.