Loss of sigma-factor of RNA polymerase of Xanthomonas campestris pv. oryzae during phage Xp10 infection

Ten min after infection of Xanthomonas campestris pv. oryzae by phage Xp10, a sharp decrease in the activity of the host RNA polymerase was observed. Host RNA polymerase from phage-infected and uninfected cells was purified, and their properties were compared. The enzyme from uninfected cells contained four polypeptides with Mr = 155,000, 155,000, 93,000, and 37,000, respectively, and assembled with a stoichiometry of alpha 2 beta beta' sigma. The enzyme from infected cells lacked the sigma-subunit. The enzyme from uninfected cells utilized Xp10 DNA and poly[d(A-T)] as templates, the enzyme from phage-infected cells failed to transcribe Xp10 DNA, but retained the ability to transcribe poly(A-T). The regions of the Xp10 genome transcribed by the two enzymes were also investigated. The enzyme from uninfected cells transcribed the leftmost 25-30% of the Xp10 genome. The enzyme from phage-infected cells also transcribed the same region, but the enzyme activity was very low. Other properties such as (a) the response to RNA polymerase inhibitors, (b) the effect of N-ethylmaleimide, (c) the requirement of Mg2+ and Mn2+, and (d) the optimum temperature and pH of the two enzymes were very similar.     

Methane production using whole and screened dairy manure in conventional and fixed-film reactors

The technical feasibility of adopting the fixed-film reactor concept for biogas production from screened dairy manure was investigated. The methane production capability of laboratory-scale 4-L anaerobic reactors (conventional and fixed-film) receiving screened dairy manure and operated at 35 degrees C was compared. Dairy manure filtrate with 4.4% total solids (TS) and 3.4% volatile solids (VS) (average value) was prepared from 1:1 manure-water slurry. The feed material was added intermittently at loading rates ranging from 2.34 to 25 and 2.25 to 785 g VS/L d, respectively, for the conventional and fixed-film reactors. Maximum methane production rate (L CH(4)/L d) for the conventional reactor was 0.63 L CH(4)/L d achieved at a 6-day hydraulic retention time (HRT). For the fixed-film reactor the maximum production rate was 3.53 L CH(4)/L d when operated at a loading rate of 262 g VS/L d (3 h HRT). The fixed-film reactor was capable of sustaining a loading of 785 g VS/L d (1 h HRT). The fixed-film reactor performed much better than the conventional reactors. These results indicate that a large reduction of required reactor volume is possible through application of a fixed-film concept combined with a liquid-solid separation pretreatment of dairy manure.     

The use of a hydroxylapatite-filter steroid receptor assay method in the study of the modulation of androgen receptor interaction

Receptors for androgen, estrogen, and glucocorticoid can be assayed by hydroxylapatite adsorption of the radioactive steroid-receptor complex and washing of the adducts on membrane filters mounted on a multiple filter holder. The method is economical, very rapid and sensitive. This new receptor assay method was used to study the modulation of androgen receptor of rat ventral prostate by metal ions, thiols, and ligand structure. The interaction of androgen with the naked receptor is inhibited by 10 microM ZnCl2, CdSO4, or CuSO4 but this inhibition is competed by androgen and is reversed by DTT. The androgen-receptor complex is less sensitive to divalent metal ions but Zn2+, at 3 mM, appears to alter the conformation of the receptor and promote the release of androgen. Certain phenanthrene derivatives exhibited striking structural specificities in their ability to compete with radioactive androgen for binding to the prostate receptor. The results suggest that the receptor has binding preference toward individual ring structure in the steroid.     

The immunological and structural comparisons of deoxyribonucleases I. Glycosylation differences between bovine pancreatic and parotid deoxyribonucleases

A rabbit antiserum against bovine pancreatic DNase A is used to study the immunological reaction of DNases I. As shown by double immunodiffusion, bovine pancreatic DNases A, B, C, and D are immunologically identical, so are DNases from bovine pancreas and parotid and from ovine pancreas. These DNases also behave similarly in immunotitration of DNase activity and all are tightly bound to the immunoaffinity medium, requiring an acidic buffer with 10% ammonium sulfate to dissociate. On the other hand, porcine pancreatic and malted barley DNases that do not form precipitin lines remain active in solution with the antibody; however, in spite of the lack of inhibition these DNases are retarded (but not tightly bound) in immunoaffinity chromatography, suggesting interaction with the antibody. In thin layer isoelectric focusing, the parotid DNase, purified with the immunoaffinity technique, shows only two major active components whose isoelectric points correspond to those of DNases A and C of bovine pancreas. As estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the molecular weight of parotid DNase is 34,000, approximately 3,000 more than that of the pancreatic enzyme. However, both parotid and pancreatic DNases have the same NH2-terminal leucine, an identical COOH-terminal amino acid sequence, nearly identical amino acid compositions, and almost the same peptide maps. The molecular weight difference is due to differences in the carbohydrate side chains. Results of peptide analyses indicate that parotid DNase contains two glycopeptides; pancreatic DNase has only one. In addition, both parotid glycopeptides contain glucosamine and galactosamine while the pancreatic glycopeptide has only glucosamine.     

Estimation of intracellular fluid volume of L cells

Dilution of ¹⁴C-sucrose solution by intracellular fluid released as a result of ultracentrifugation was used to estimate the intracellular fluid volume of L cells. Consistent relationships to total cell volume as estimated by use of an electronic particle counter were obtained. Expressed as a percentage of total cell volume, the mean value plus or minus the S.D. for 6 experiments was 72.8 ± 0.9.     

受白粉菌诱导大麦抗感等基因系蛋白质变化的双向电泳分析

分别对接种与否的大麦抗—感白粉病等基因系—叶期幼苗取材进行蛋白质双向电泳分析。结果表明,病原的侵入使抗—感两系在30Kd以下的低分子量区域的蛋白质发生了明显变化。接种48小时之后,抗病系在pH5.5、6.0、6.8及8.8附近出现了对照中所没有的蛋白质,而在pH6.0和8.8附近的蛋白质则较对照有减小的趋势;感病系在pH6.0附近蛋白质明显增多,在pH8.8处不仅在量上有大幅度提高,而且种类也有增加。结果还表明,抗—感系间在未接种的情况下双向电泳图谱也有差异,接种之后由于感病系在pH8.8处蛋白质的特异性合成,使抗—感两系间的差异缩小。