Enzyme assay in cultures of Escherichia coli by a continuous flow method based on lysis from without by a phage ghost.

Lysis of Escherichia coli from without by excess of phage ghost has been shown to give excellent yield of several enzymes. The application of lysis from without to a continuous determination of enzymes in growing cultures of E. coli is illustrated with β-galactosidase. This application can be used in studies on changes of a large number of enzymes during metabolic perturbation (induction, repression, etc.) of growing cultures of E. coli.     

Endocrine steroid sulfotransferases: steroid alcohol sulfotransferase from human breast carcinoma cell line MCF-7

Steroid alcohol sulfotransferase (SAS) has been isolated from the cytosol of a human breast carcinoma cell line, MCF-7. This enzyme from Sephadex G-200 chromatography displayed a mol. wt of 118 KDa. The conditions for optimal enzymic activity of SAS were determined to be 20 min incubations at 45 degrees C in 0.2 M Tris buffer (pH 7.5) containing 0.06 M Mg2+. Chromatofocusing chromatography also yielded a single peak of SAS with a pI of 5.8. Results from the incubations of a series of androstane analogues revealed that SAS required a 3 beta-hydroxyl on a steroid with the trans bridge between the A and B rings. Neither the 3 beta-allylic hydroxyl group nor the A-ring phenolic 3-hydroxyl accepted the sulfate group from 3'-phosphoadenosine-5'-phosphosulfate. D-ring beta-hydroxyl groups were tolerated by the enzyme, however, alpha-hydroxyl groups on the D-ring appeared to interfere with the reaction. Sulfurylation of steroids by SAS was related inversely to the sum of the displacements of the 3-hydroxyl plus that of the 17-hydroxyl groups relative to the plane of symmetry of the dehydroepiandrosterone nucleus. This enzyme was also capable of sulfurylating short chain aliphatic alcohols, although at greatly reduced rates. 3 beta-Chloro-5-androstene-17-one and 2-nitroestradiol. 17 beta proved to be the best inhibitors of SAS.     

Evaluation of the 2‐(1‐Hexyloxyethyl)‐2‐devinyl pyropheophorbide (HPPH) mediated photodynamic therapy by macroscopic singlet oxygen modeling [J. Biophotonics 9, No. 11–12, 1344–1354 (2016)]

In the article by R. Penjweini, M. M. Kim et al. (doi: 10.1002/jbio.201600121 ), published in J. Biophotonics 9, 1344–1354 (2016), the constants C₀₁, C₀₂, b₁, and b₂ determined from fitting the fluorescence single value decomposition (SVD) for phantoms with different optical properties and the corresponding Figure 2(a) are not correct. This erratum is published to correct the Section 2.3 and Figure 2(a).     

Plasma membrane-associated cysteine proteinases in human and animal tumors

The ability of tumor cells to invade into and through normal tissue during the metastatic cascade has been attributed to tumor-associated degradative enzymes including proteinases of the metallo, serine and cysteine classes. Work from several laboratories has established that the cysteine proteinases cathepsins L and B are released from tumor cells, primarily as latent precursor forms. In addition, a cathepsin B-like cysteine proteinase has been shown to be associated with the plasma membrane fraction of several animal and human tumors. This form of the enzyme retains activity under physiologic (or pathologic) conditions including at neutral pH and in the presence of low Mr inhibitors. Since we have established that cathepsin B can degrade the basement membrane attachment glycoprotein laminin, we speculate that plasma membrane-associated cathepsin B may participate in focal dissolution of the basement membrane during tumor cell extravasation.     

Transport and accumulation of PVP-Hypericin in cancer and normal cells characterized by image correlation spectroscopy techniques

PVP-Hypericin (PVP: polyvinylpyrrolidone) is a potent anti-cancer photosensitizer for photodynamic diagnosis (PDD) and therapy (PDT). However, cellular targets and mechanisms involved in the cancer-selectivity of the photosensitizer are not yet fully understood. This paper gives new insights into the differential transport and localization of PVP-Hypericin in cancer and normal cells which are essential to unravel the mechanisms of action and cancer-selectivity. Temporal (TICS) and spatiotemporal (STICS) image correlation spectroscopy are used for the assessment of PVP-Hypericin diffusion and/or velocity in the case of concerted flow in human cervical epithelial HeLa and human lung carcinoma A549 cells, as well as in human primary dendritic cells (DC) and human peripheral blood mononuclear cells (PBMC). Spatiotemporal image cross-correlation spectroscopy (STICCS) based on organelle specific fluorescent labeling is employed to study the accumulation of the photosensitizer in nucleus, mitochondria, early-endosomes and lysosomes of the cells and to assess the dynamics of co-migrating molecules. Whereas STICS and TICS did not show a remarkable difference between the dynamics of PVP-Hypericin in HeLa, A549 and DC cells, a significantly different diffusion rate of the photosensitizer was measured in PBMC. STICCS detected a stationary accumulation of PVP-Hypericin within the nucleus, mitochondria, early endosomes and lysosomes of HeLa and A549 cells. However, significant flow due to the directed motion of the organelles was detected. In contrast, no accumulation in the nucleus and mitochondria of DC and PBMC could be monitored.     

Membrane-associated cathepsin L: a role in metastasis of melanomas

Subcellular distribution of cathepsin L, the major protein released by transformed or ras transfected fibroblasts, was examined in murine liver, murine B16 amelanotic melanoma and human A2058 melanoma after sequential differential and Percoll density gradient centrifugation. In both murine and human melanomas, cathepsin L activity was found to be enriched in plasma membrane fractions; cathepsin L in these fractions was in both native and acid activatable forms. Plasma membrane fractions from B16 melanoma subpopulations of "low" and "high" metastatic potential were assayed for activity of cathepsin L and of heat stable endogenous inhibitors. The relative specific activity of cathepsin L was 7-fold greater in the subpopulation of "high" metastatic potential, whereas cysteine proteinase inhibitory activity was 5-fold less. Since cathepsin L can degrade intact basement membrane, this membrane-associated cathepsin L may well contribute to metastatic spread of melanomas.     

Demographic parameters of greenbug,Schizaphis graminum (Rondani) (Hemiptera: Aphididae), on six Iranian genotypes of barley

Greenbug, Schizaphis graminum (Rondani), is one of the injurious aphids of cereals in various regions of the world. This study has measured the life table parameters of the greenbug on six barley genotypes at 25 ± 2 °C, 55 ± 10% RH and 16:8 L:D in greenhouse. According to the results, significant differences were not observed for aphids’ developmental times among the genotypes. Also, the nymphs underwent no mortality on any of the tested genotypes. The longevity of the aphids was obtained from 23.7 to 35.9 days. The least mean number of offsprings was on Raihan cultivar and the highest on line13 (Legia/CWB117-5-9-5). R₀ value was significantly higher on line 20 (Mall-4-3094-2//Alpha/Cum/3/Victoria/ICB01-1368-0AP) and line 13 than on the Raihan cultivar. However, the r_m and λ values were significantly higher on line 44 (Sls/Bda//Sararood-1) than on Raihan cultivar. T (mean generation time) and DT (doubling time) values of the Raihan cultivar were longer than the other genotypes. Results of this research indicated that among the tested genotypes, the Raihan cultivar is the most unsuitable host for greenbug aphid and lead to the decrease of greenbug population growth.     

Studies on bovine adrenal estrogen sulfotransferase. III. Facile synthesis of 3'-phospho- and 2'-phosphoadenosine 5'-phosphosulfate.

A practical synthesis of 3'-phosphoadenosine 5'-phosphosulfate (IV) in yields of 68-72% from adenosine 2',3'-cyclic phosphate 5'-phosphate (II) is described. Reaction of II with triethylamine-N-sulfonic acid affords adenosine 2',3'-cyclic phosphate 5'-phosphosulfate (III) which, on treatment with ribonuclease-T2, provides IV. Spleen phosphodiesterase, on the other hand, converts III to 2'-phosphoadenosine 5'-phosphosulfate (V). The biological activity of IV, measured by sulfate transfer to [6,7-3H2]estrone as mediated by bovine adrenal estrone sulfotransferase (3'-phosphoadenylyl-sulfate:estrone 3-sulfotransferase, EC 2.8.2.4), is identical with that obtained with a sample of IV prepared by an established biochemical procedure. By contrast, V exhibits approximately one-third the activity of the natural isomer.