Secretion of the sweet-tasting plant protein thaumatin byBacillus subtilis

Summary With the -amylase promoter and ribosome binding site,Bacillis subtilis was used to express the sweet plant protein thaumatin II cDNA fused in the correct reading frame to the -amylase leader peptide. The r-thaumatin was purified from the medium on a S-Sepharose column and detected with western blots by sheep -thaumatin antibodies. The r-thaumatin and authentic thaumatin were the same size when reduced by 2-ME and the same size when not reduced.     

User requirements on CT-based computed dose planning systems in radiation therapy. Presentation of 'check lists'

The expanding use of computers in radiation therapy procedures, especially the rapidly increasing use of digital CT-information, necessitates the coordination of the different systems in order to facilitate their developments. In order to define necessary demands for tomorrow a Nordic cooperation was initiated 1981 by NORDFORSK (Nordic co-operative organization for applied research), and a group of physicians and physicists having their daily work in this field of medicine and physics was invited to produce a report on 'User requirements on CT-based computed dose planning systems on radiation therapy'. The work has been done within the frame of NORDFORSK's activities and has been independent of the existing commissions and associations in the radiology field, but it has taken into consideration recommendations that have been given by or are being produced by other organizations. This report is a short summary of the complete paper which will be published in Acta Radiologica. The aim of this short version is to get an early presentation of the 'requirement lists' (see Appendix) which we think are of immediate importance.     

Secretion of the sweet-tasting plant protein thaumatin byStreptomyces lividans

Summary To produce and direct the export inStreptomyces lividans of the sweet plant protein thaumatin, thaumatin II cDNA was fused in the correct reading frame to the -galactosidase leader peptide, under the control of the -galactosidase promoter and ribosome binding site. The export of the recombinant thaumatin may allow the correct formation of the thaumatin disulfide bonds. The recombinant thaumatin was purified from the medium on an S-Sepharose column and detected with western blots by sheep -thaumatin antibodies. The recombinant thaumatin was the same size as authentic thaumatin and changed position on an acrylamide gel in response to reduction by 2-mercaptoethanol in the same manner.     

Oligochaetes and water pollution in two deep Norwegian lakes

Analyses of the oligochaete fauna of two of the deepest lakes in Scandinavia — the Norwegian lakes Mjösa (450 m) and Tyrifjorden (295 m), revealed a totally different species composition in the deep profundal compared with the upper profundal - in contact with the nutrient-enriched epilimnion. In both lakes a pronounced thermal stratification develops in the summer, thus the epilimnion receiving gross organic pollution behaves differently from the profundal. The lakes are each effectively divided into two bodies of water with limited water exchange between them, i.e. one major oligotrophic body and one minor more nutrient-rich. Since the 1950s both lakes have been exposed to heavy pollution of various kinds. In Lake Mjösa in 1975 and 1976 unpleasant algal blooms of the blue-green alga Oscillatoria bornetii fa. tenuis occurred. Bottom samples obtained at the same time revealed that the deep central bottoms of the lake were totally dominated by oligotrophic oligochaete indicators, i.e. by Stylodrilus heringianus and Spirosperma ferox, while the fauna of the upper profundal in the vicinity of domestic and agricultural sewage outfalls, wood processing industries, etc. was dominated by Limnodrilus hoffmeisteri and Tubifex tubifex in great abundance, indicating enriched conditions. Several other species indicative of eutrophy, were absent, most of them belonging to the genus Potamothrix. A fairly similar situation exists in Lake Tyrifjorden, where, for instance, in the shallow bay of Steinsfjorden — heavily eutrophied by agricultural wastes — blooms of blue-green algae have caused problems from time to time. The same oligochaete communities as in Lake Mjösa distinguish the central oligotrophic bottoms from the regionally more enriched upper profundal. The likely reasons for an intact profundal oligochaete fauna are great volumes of oxygen-rich hypolimnic water of low temperature and a high bottom/lake surface area ratio.     

Flow cytometric multiparameter analysis of proliferating cell nuclear antigen/cyclin and Ki-67 antigen: a new view of the cell cycle

Flow cytometric multiparameter analysis of two proliferation-associated nuclear antigens (proliferating cell nuclear antigen (PCNA)/cyclin and Ki-67) was performed on seven human hematopoietic cell lines. PCNA/cyclin, an S phase-related antigen, was detected using an autoantibody and a fluorescein isothiocyanate-labeled anti-human antibody. The Ki-67 antigen, which in cycling cells is expressed with increasing levels during the S phase with a maximum in the M phase, was detected using a monoclonal antibody and a phycoerythrin-conjugated anti-mouse antibody. In some experiments the PCNA/Ki-67 staining was combined with a DNA stain, 7-amino actinomycin D, and simultaneous detection of the three stains was performed by a single laser flow cytometer. Using this technique four distinct cell populations, representing G1, S, G2, and M, respectively, could be demonstrated in cycling cells on the basis of their PCNA/cyclin and Ki-67 levels. The cell cycle phase specificity could be verified using metaphase (vinblastine, colcemide) and G2 phase (mitoxantrone) blocking agents, as well as by stainings with a mitosis-specific antibody (MPM-2). Also, G0 cells could be discriminated from G1 cells in analysis of a mixture of resting peripheral mononuclear blood cells and a proliferating cell line. This technique can be valuable in detailed cell cycle analysis, since all cell cycle phases can be visualized and calculated using a simple double staining procedure.     

Complement activation,regulation, and molecular basis for complement‐related diseases

The complement system is an essential element of the innate immune response that becomes activated upon recognition of molecular patterns associated with microorganisms, abnormal host cells, and modified molecules in the extracellular environment. The resulting proteolytic cascade tags the complement activator for elimination and elicits a pro‐inflammatory response leading to recruitment and activation of immune cells from both the innate and adaptive branches of the immune system. Through these activities, complement functions in the first line of defense against pathogens but also contributes significantly to the maintenance of homeostasis and prevention of autoimmunity. Activation of complement and the subsequent biological responses occur primarily in the extracellular environment. However, recent studies have demonstrated autocrine signaling by complement activation in intracellular vesicles, while the presence of a cytoplasmic receptor serves to detect complement‐opsonized intracellular pathogens. Furthermore, breakthroughs in both functional and structural studies now make it possible to describe many of the intricate molecular mechanisms underlying complement activation and the subsequent downstream events, as well as its cross talk with, for example, signaling pathways, the coagulation system, and adaptive immunity. We present an integrated and updated view of complement based on structural and functional data and describe the new roles attributed to complement. Finally, we discuss how the structural and mechanistic understanding of the complement system rationalizes the genetic defects conferring uncontrolled activation or other undesirable effects of complement.     

Structural basis of the lisinopril-binding specificity in N- and C-domains of human somatic ACE

Angiotensin I-converting enzyme (ACE) is a dipeptidyl carboxypeptidase which converts angiotensin I into the vasopressor peptide angiotensin II and also inactivates the hypotensive peptide bradykinin, playing an important role in blood pressure regulation. The present work describes the molecular modeling of the N-terminal human somatic ACE in complex with the inhibitor lisinopril, identifying the residues involved in the inhibitor-binding pocket. The obtained results identify differences in the lisinopril lysine moiety-binding residues for N- and C-terminals of sACE domains and an important carboxy-terminal proline hydrophobic accommodations mediated by the aromatic ring of Tyr532 and Tyr1128 residues, respectively. The present model will be useful for the development of a new inhibitor family based on the natural BPP peptides and derivatives, or even to improve the binding capacities and the domain specificity of the already known inhibitors.