Cellular thorn-like projections in human endocervical epithelium. Preliminary scanning electron-microscopic study.

By means of scanning electron-microscopic investigations, thorn-like projections (TLP) were observed in the cilio-secretory epithelium of the ventral surface of the human endocervix. These projections, which seem to be characteristic of a new cell type, were seen at different stages of the menstrual cycle (days 8, 14 and 21) in 4 apparently healthy fertile women. Striking differences in length, with a maximum size at midcycle, suggest an evolutive process throughout the menstrual cycle. The origin and possible physiological role of TLP in the reproductive process are discussed.     

The effect of pH on the oxygen kinetics of cytochrome c oxidase proteoliposomes

The effect of pH on the oxygen kinetics of cytochrome c oxidase incorporated into phospholipid vesicles is studied. The pH profiles of the oxygen kinetics of energized and deenergized oxidase vesicles are similar. An effect of pH on the slope of the reciprocal plot of rate against oxygen concentration is observed, and this may indicate that protons are involved in the rate limiting step of the reaction between oxygen and reduced oxidase. In contrast to the pH dependence of the oxygen kinetics, the binding of CO to the oxidase is not pH dependent.     

Partial N-terminal amino acid sequence of pro-opio-melanocortin (ACTH/beta-LPH precursor) from rat pars intermedia

Posterior lobes of rat pituitary (pars intermedia plus pars nervosa) were incubated with various labeled amino acids and the cell extracts analyzed by NaDodSO₄ polyacrylamide disc gel electrophoresis. Two forms of precursor proteins for betaendorphin and alpha-MSH were synthesized. Both forms have been shown to contain the fragments beta-LPH 61–69 and ACTH 1–8 and are thought to have the same peptide backbone. The two forms were simultaneously submitted to automatic Edman degradation and the following partial sequence was obtained: Trp/Arg₁-Leu₃-Phe₄-Ser₅-Ser₆-Leu₁₁-Thr_12–13-Tyr₁₄-Ser₁₅-Leu_17–18-Ala₁₉-lle₂₁-Arg_22–25- Leu_26–28-Ser₂₉. This sequence was compared with that reported by Nakanishi et al. (18). Their amino acids sequence was indirectly derived from DNA sequencing after isolation of mRNA from bovine pars intermedia. This comparison indicates the presence of a signal peptide of 26 amino acids in the sequence of beef ACTH/beta-LPH precursor.     

Biosynthesis and characterization of adrenocorticotropic hormone, alpha-melanocyte-stimulating hormone, and an NH2-terminal fragment of the adrenocorticotropic hormone/beta-lipotropin precursor from rat pars intermedia.

Isolated intermediate lobe cells from 40 rat pituitaries were incubated for 3 h with [35S]methionine + [3H]-phenylalanine, [35S]methionine, [3H]valine, and [3H]leucine. The cell extracts were purified by carboxymethyl-cellulose chromatography (CMC) and the fraction eluting with ovine adrenocorticotropic hormone (ACTH) was further purified either by another CMC under the same conditions or by high performance liquid chromatography (HPLC). Microsequencing of the product from the second CMC allowed the identification of a peptide containing methionine 4 and phenylalanine 7, as expected for the NH2 terminus of ACTH. Purification by HPLC of a similar peptide obtained from the three other incubations gave three main raoactive peaks which were further characterized by their migration rates on polyacrylamide gels, molecular weight, and microsequencing. Results indicated that intact ACTH (residues 1-39) is present in extracts of rat intermediate lobe, but in very small quantities (less than 1% of the beta-endorphin content). ACTH is probably broken down into smaller fragments, e.g. alpha-melanocyte-stimulating hormone (alpha-MSH) (ACTH, 1-13) and corticotropin-like intermediate lobe peptide (CLIP) (ACTH, 18-39). These studies also revealed with existence of a peptide having identical sequence with the (N-1) terminus of the ACTH/lipotropin (LPH) precursor.     

Histochemical studies on the oil-bodies ofMarchantia paleacea bert

Summary Correlated light and electron microscope histochemistry was performed on the oil bodies (OB) ofMarchantia paleacea. It was revealed that they contain significant quantities of material positive to PAS and periodic acid-thiocarbohydrazide-silver proteinate (PA-TCH-SP) reactions. Ag deposits were localized on the matrix surrounding the globules and on the distinct one close to the inner face of the OB membrane. In addition, the lipophilic globules were peripherally impregnated with staining product. Polysaccharidic material appears to be elaborated in dictyosomes and then advanced into the OB by their vesicles.The OB show a lipophilic character and are intensely stained with Sudan black B, with an aqueous solution of osmium tetroxide and with the Nadi reaction as modified byDavid andCarde (1964).The performance of mercury bromophenol blue, aniline blue black, coomassie brilliant blue R and acid fuchsin stainings showed that the OB do not contain structurally detectable proteins. Besides, a positive reaction of the inner OB was obtained in fresh tissue with a variety of tests sensitive to phenolic or other aromatic ring compounds. On the contrary, the epidermal and scale OB containing small globules are obviously stained only with Gibbs' reagent. The colours obtained with the above reactions, were almost the same,i.e. brown with a red shade, except for the blue one of the Gibbs' reagent.The presented observations favour the conclusion that the OB matrix does not contain proteins, but substances polysaccharidic in nature. Phenolic and possibly other aromatic compounds appear to be present at least in inner OB.     

Primary sequence of the glucanase gene from Oerskovia xanthineolytica. Expression and purification of the enzyme from Escherichia coli

A 2.7-kilobase fragment of DNA from Oerskovia xanthineolytica containing the gene for a beta-1,3-glucanase has been isolated and its complete nucleotide sequence determined. The sequence was found to contain two large open reading frames. Purification of the mature native enzyme and subsequent amino-terminal sequencing defined the glucanase gene in one reading frame which potentially encodes a protein of 548 amino acids. We have expressed this glucanase gene in Escherichia coli under control of the lacUV5 promoter and found the product to be secreted into the periplasm as a mature enzyme of about the same molecular weight as that of the native protein. The recombinant enzyme was purified to near homogeneity by a single step of high performance liquid chromatography. The ability of the recombinant enzyme to digest beta-glucan substrates and to lyse viable yeast cells was found to be indistinguishable from that of the native protein. Deletion of the cysteine-rich carboxyl-terminal 117 amino acids of the enzyme, which also contain two duplicated segments, abolished the lytic activity but did not significantly affect the glucanase function of the protein. The possible involvement of this domain in interaction with the yeast cell wall is discussed.     

Mineralization of 2,4-dichlorophenoxyacetic acid (2,4-D) in soil inoculated with Pseudomonas cepacia DBO1(pRO101), Alcaligenes eutrophus AEO106(pRO101) and Alcaligenes eutrophus JMP134(pJP4): effects of inoculation level and substrate concentration

Mineralization of 2,4-dichlorophenoxyacetic acid (2,4-D) by two Alcaligenes eutrophus strains and one Pseudomonas cepacia strain containing the 2,4-D degrading plasmids pJP4 or pRO101 (=pJP4::Tn1721) was tested in 50 g (wet wt) samples of non-sterile soil. Mineralization was measured as ¹⁴C-CO₂evolved during degradation of uniformly-ring-labelled ¹⁴C-2,4-D. When the strains were inoculated to a level of approximately 10⁸ CFU/g soil, between 20 and 45% of the added 2,4-D (0.05 ppm, 10 ppm or 500 ppm) was mineralized within 72 h. Mineralization of 0.05 ppm and 10 ppm, 2,4-D by the two A. eutrophus strains was identical and rapid whereas mineralization by P. cepacia DBO1(pRO101) occurred more slowly. In contrast, mineralization of 500 ppm 2,4-D by the two A. eutrophus strains was very slow whereas mineralization by P. cepacia DBO1 was more rapid. Comparison of 2,4-D mineralization at different levels of inoculation with P. cepacia DBO1(pRO101) (6×10⁴, 6×10⁶ and 1×10⁸ CFU/g soil) revealed that the maximum mineralization rate was reached earlier with the high inoculation levels than with the low level. The kinetics of mineralization were evaluated by nonlinear regression analysis using five different models. The linear or the logarithmic form of a three-half-order model were found to be the most appropriate models for describing 2,4-D mineralization in soil. In the cases in which the logarithmic form of the three-half-order model was the most appropriate model we found, in accordance with the assumptions of the model, a significant growth of the inoculated strains.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - CFU colony forming units - PTYG peptone, tryptone, yeast & glucose - DPM disintegrations per minute     

Isolation and partial characterization of a biosynthetic N-terminal methionyl peptide of bovine pars intermedia: relationship to ubiquitin

During in vitro labelling studies of beef pituitary intermediate lobe cells, a 4000 daltons molecular weight peptide was found to be biosynthesized in major yields. The partial amino acid sequence of this peptide has been found to be Met 1, Leu 8, 15 and Lys 6, 11, 27, 29, 33. This partial sequence fits very well the one expected from the N-terminal sequence of bovine and human ubiquitin, a non-histone fragment of the nuclear protein A24. Since an identical peptide was also biosynthesized from rat hypothalamus and mouse AtT-20 tumor cells, the ubiquitous nature of this peptide is further revealed.