A Nucleoside Diphosphate Kinase from Paramecium tetraurelia with Protein Kinase Activity

Nucleoside diphosphate kinase (NDP kinase) from Paramecium was purified to homogeneity. The native enzyme was 80 kDa (by gel filtration), with subunits of 18 and 20 kDa. Near the amino terminus, 15 of 20 residues were identical with those in human NDP kinase, and 17 of 20 with the awd gene product from Drosophila. NDP kinase bound α-labeled ATP and GTP, and a photoreactive GTP analog labeled both subunits. Purified NDP kinase underwent autophosphorylation on a histidine and a serine residue using either ATP or GTP as a substrate. The enzyme also catalyzed acid-stable phosphorylation of casein and phosvitin. This protein kinase activity is distinct from the histidine phosphorylation that is part of the NDP kinase catalytic cycle. Antiserum against the purified protein from Paramecium cross-reacted with 16- to 20-kDa proteins in most species tested, and with a larger protein (44 kDa) in Paramecium, Xenopus, and two human lines. The multiple forms (20 and 44 kDa) of the NDP kinase in Paramecium and its protein kinase activity, suggest that the protein is more than a housekeeping enzyme; it may have regulatory roles such as those of the NDP kinase-like awd protein of Drosophila and Nm23 protein of humans.     

The Total Chemical Synthesis of Monocyte Chemotactic Protein-1 (MCP-1)

The affinity-based N^α-amino protecting group tetrabenzo [a,c,g,i]fluorenyl-17-methoxycarbonyl (Tbfmoc) has been utilized as a hydrophobic probe to allow the simple, quick and highly effective isolation of a 76 residue cysteine-containing protein (MCP-1). The base-labile Tbfmoc group can be removed under very mild conditions, which preserve the thiol-con taining protein in the reduced state. Oxidative folding was then used to furnish the biologically active β-chemokine MCP-1.     

CD23McAb抑制活化B细胞增殖机制的初步探讨

本研究用ＣＤ２３单克隆抗体交联活化的人扁桃体Ｂ细胞,证明ＣＤ２３ＭｃＡｂ对Ｂ细胞呈双向调节效应,即：高浓度区抑制Ｂ细胞增殖,低浓度区促进Ｂ细胞增殖,继而,用对Ｂ细胞有抑制效应浓度的ＣＤ２３ＭｃＡｂ交联Ｂ细胞膜ＣＤ２３分子,通过研究抑制效应恢复条件,探讨了ＣＤ２３ＭｃＡｂ产生抑制效应的作用机制。结果显示：去除ＣＤ２３ＭｃＡｂ后,抑制作用不能恢复,用ＳＡＣ再次活化,使Ｂ细胞恢复增殖状态,用促Ｂ细胞增殖     

Distribution and Subcellular Localization of High-Molecular-Weight Microtubule-Associated Protein-2 Expressing Exon 8 in Brain and Spinal Cord

Abstract: The expression of high-molecular-weight (HMW) microtubule-associated protein-2 (MAP-2) expressing exon 8 (MAP-2+8) was examined by immunoblotting during rat brain development and in sections of human CNS. In rat brain, HMW MAP-2+8 expression was detected at embryonic day 21 and increased during postnatal development. In adult rats, HMW MAP-2+8 comigrated with MAP-2a. In human adult brain, HMW MAP-2+8 was expressed in select neuronal populations, including pyramidal neurons of layers III and V of the neocortex and parahippocampal cortex, pyramidal neurons in the endplate, CA2 and subiculum of the hippocampus, and the medium-sized neurons of the basal ganglia. In the cerebellum, a subpopulation of Golgi neurons in the internal granular cell layer and most Purkinje cells were also stained. In the spinal cord staining was observed in large neurons of the anterior horn. Staining was present in cell bodies and dendrites but not in axons. At the ultra-structural level, HMW MAP-2+8 immunoreactivity was observed on mitochondrial membranes and in postsynaptic densities (PSDs) of some asymmetric synapses in the midfrontal cortex and spinal cord. Immunoblots of proteins isolated from enriched mitochondrial and PSD fractions from adult human frontal lobe and rat brains confirmed the presence of HMW MAP-2+8. The presence of HMW MAP-2+8 in dendrites and in close proximity to PSDs supports a role in structural and functional attributes of select excitatory CNS synapses.     

Chemical induction of interleukin-8, a proinflammatory chemokine,in human epidermal keratinocyte cultures and its relation to cytogenetic toxicity

Tumor promoters, proinflammatory cytokines, endotoxins, and protein synthesis inhibitors can modulate cell cycle kinetics of various cell types, stimulate production of reactive oxygen species, and induce keratinocytes to produce interleukin-8 (IL-8), a potent chemotactant for polymorphonuclear neutrophils and T lymphocytes. The aim of this study was to determine whether perturbations of cytogenetic responses correlated with the induction of IL-8 expression. Cultures of primary human keratinocytes were grown in serum-free medium with 5 mol/L bromodeoxyuridine to label DNA and exposed either to phorbol-13-myristate-12-acetate (PMA) (0.0001–100 ng/ml), cycloheximice (CHX) (0.01–50 g), lipopolysaccharide (0.1–100 g/ml), tumor necrosis factor- (TNF) (3.13–50 ng/ml), or interleukin-1 (IL-1) (1–182 pg/ml). Metaphase chromosome preparations were stained by a fluorescence-plus-Giemsa technique to differentiate sister chromatids. For IL-8 production, keratinocytes were grown to 70% confluency and then exposed to chemicals for 24 h. Immunoreactive IL-8 was quantitated from the supernatants by ELISA. With the exception of benzo(a)pyrene used as a positive control, none of the agents induced sister chromatid exchanges. However, PMA and TNF induced IL-8 production that coincided with significant cell cycle inhibition. IL-1 had no effect on cytogenetic endpoints, yet stimulated a 6.3-fold increase in IL-8. CHX inhibited cell cycle progression and mitotic activity at concentrations that were 200 times lower than required for IL-8 induction; however, puromycin (0.31–10 g/ml), another protein synthesis inhibitor, did not induce IL-8. At all concentrations tested, TNF reduced the mitotic index by 45%, slowed cell cycle progression by 3.5 h, and induced a flat, albeit large, IL-8 response at concentrations 12.5 ng/ml. These agent-specific response patterns suggest that induction of IL-8 production is not always the inevitable result of cell cycle perturbations or genetic damage.Abbreviations B(a)P benzo(a)pyrene - BrdU 5-bromo-2-deoxyuridine - CHX cycloheximide - ICAM intercellular adhesion molecules - IL-1 interleukin-1 - IL-8 interleukin-8 - KGM keratinocyte growth medium - LPS lipopolysaccharide - PKC protein kinase C - PMA phorbol-13-myristate-12-acetate - PMN polymorphonuclear neutrophil - ROS reactive oxygen species - SCE sister chromatid exchange - TNF tumor necrosis factor      

Influence of carbon and nitrogen sources on glutathione catabolic enzymes inCandida albicans during dimorphism

The effect of carbon sources, glucose and sucrose, and nitrogen sources such as ammonia, glutamate andl-citrulline on the activities of glutathione metabolic enzymes has been studied. Yeast and mycelial cells were used to identify changes in activity levels of glutathione reductase (GSSGR), glutathione transferase (GST), glutathione peroxidase (GPX) and -glutamyl transpeptidase (GGT). Enzyme activities from cells grown in sucrose media were lower than in glucose media regardless of the enzyme tested, morphological form, or the growth interval. In all enzymes except GST, activity was higher in yeast form than in mycelia, regardless of nitrogen source, with lower activity from 24 to 72 h than at 96 h. In citrulline media, yeast form showed the maximum GST, GGT, and GPX activity. In ammonia-amended media, mycelia showed maximum activity in GGT, whereas in glutamate media, mycelia showed the maximum activity in GST. Also, the type of nitrogen source had no effect on GPX activity in the mycelial form. Finally, changing the nitrogen source showed no significant effect on GSSGR activity, either in the yeast or mycelial form.     

Pharmacokinetics of anti-ganglioside GD2 mAb 14G2a in a phase I trial in pediatric cancer patients

A phase I trial of a murine anti-ganglioside (GD2) monoclonal antibody (mAb) 14G2a was conducted in 14 neuroblastoma patients and 1 osteosarcoma patient to assess its safety, toxicity and pharmacokinetics in pediatric patients. The pharmacokinetics of mAb 14G2a were biphasic with at ₁ ^2/ of 2.8±2.8 h and at ₁ ^2/ of 18.3±11.8 h. In general,t ₁ ^2/ was dose-dependent with a level of significance ofP=0.036, and it reached a plateau at doses of 250 mg/m² or more. Overall the peak serum levels were dose-dependent atP<0.001. However, they demonstrated an abrupt increase between doses of 100 mg/m² and 250 mg/m². The latter two suggest a saturable mechanism for mAb elimination. In addition, peak serum concentrations were observed earlier at higher mAb doses, which indicates the achievement of a steady state. Thet ₁ ^2/ of mAb 14G2a in children appears to be shorter than in adults. Furthermore, 2 patients demonstrated a considerable decrease int ₁ ^2/ following retreatment with 14G2a. This was paralleled by high human anti-(mouse Ig) antibody levels. This study represents the first comprehensive analysis of murine mAb pharmacokinetics in children and will be useful in the future design of mAb therapy.This work was supported by grants from FDA, FD-R-000377 and NIH U10 CA 28439 and in part by a grant from the general Clinical Research Center program, MOI RR00827, of the National Center for Research Resources, National Institutes of Health. M. M. U.-F. and C.-S. H. were supported in part by a grant from the Children's Cancer Research Foundation, and R. A. R. was supported in part by NIH grant CA 42508     

Expression domains of the Cf1a POU domain protein during Drosophila development

We have used antibodies directed against a unique portion of the Drosophila POU domain protein Cfla to localize its sites of expression in developing embryos. Cfla protein is first detected during germ band extension in the tracheal placodes and in the midline mesectoderm cells. Tracheal expression continues throughout embryonic development, especially in the main longitudinal tracheal trunks. Additional sites of high Cfla expression are in the anterior portion of the hindgut, the roof of the stomodeum, a subset of central nervous system cells, the oenocytes, and the ring gland. In addition, Cfla expression was localized in embryos mutant for several loci involved in determining fate along the midline of the CNS and the tracheal system. Cfla midline cell expression is dependent on proper single-minded gene function, and Cfla either regulates or acts in parallel to the genes pointed and rhomboid during midline CNS and tracheal development.     

苯并［a］芘在土壤－水－水稻系统中传输动力学研究

建立了ＢａＰ在土壤水稻系统中传输模型,并对其在系统各组分中的含量变化做了定量分析。实验求得了ＢａＰ在系统各相间传输速度常数和分配系数。揭示了水中ＢａＰ向水稻体传输路径和它们的速度大小。水稻从水中吸收ＢａＰ的速度比其从土壤中吸收的速度高４倍,水中ＢａＰ向土壤沉积速度比水稻从水中吸收ＢａＰ的速度高７倍。水中ＢａＰ在迁移到达水稻体之前,绝大部分被土壤牢牢吸附而很难被水稻吸收。     

Photodynamic herbicides: 1. Concept and phenomenology

A new approach to the design of conceptually and phenomenologically new herbicides is described. It involves the joint utilization of tetrapyrrole precursors, such as δ-aminolaevulinic acid (a biodegradable amino acid) and activators of the chlorophyll biosynthetic pathway, such as 2,2′-dipyridyl, in order to induce treated plants to biosynthesize and accumulate massive amounts of tetrapyrrole intermediates of the chlorophyll biosynthetic pathway in the dark (i.e. at night). During the subsequent light period (daylight) the accumulated tetrapyrroles act as potent photodynamic sensitiziers, which in turn result in the death of susceptible plants in a matter of hours. We have therefore proposed to name herbicides that act via this mechanism as photodynamic herbicides, or more pictorially as laser herbicides. From a limited survey of agricultural plant and weed species it appears that photodynamic herbicides exhibit a very pronounced organ, age and species-dependent selectivity. For example, dicotyledonous weeds such as mustard, red-root pigweed, common purslane and lambsquarter are very susceptible while monocotyledonous plants such as corn, wheat, barley and oats are not. The biochemical basis of this selectivity seems to lie, among other things, in the rates of tetrapyrrole turnover and in a differential enhancement by the applied chemicals of the monovinyl and divinyl tetrapyrrole biosynthetic pathways in the various species. A survey of various groups of chemicals (herbicides and other selected biochemicals) that are likely to exhibit photodynamic herbicidal properties is currently under investigation.