In vivo protein cyclization promoted by a circularly permuted Synechocystis sp PCC6803 DnaB mini-intein

A synthetic Synechocystis sp. PCC6803 DnaB split mini-intein gene was constructed for the in vivo cyclization of recombinant proteins expressed in Escherichia coli. The system was used to cyclize the NH2-terminal domain of E. coli DnaB, the structure of which had been determined previously by NMR spectroscopy. Cyclization was found to proceed efficiently, with little accumulation of precursor, and the product was purified in high yield. The solution structure of cyclic DnaB-N is not significantly different from that of linear DnaB-N and it unfolds reversibly at temperatures similar to14 degreesC higher. Improved hydrogen bonding was observed in the first and last helices, and the length of the last helix was increased, while the 9-amino acid linker used to join the NH2 and COOH termini was found to be highly mobile. The measured thermodynamic stabilization of the structure (DeltaDeltaG approximate to 2 kcal/mol) agrees well with the value estimated from the reduced conformational entropy in the unfolded form. Simple polymer theory can be used to predict likely free energy changes resulting from protein cyclization and how the stabilization depends on the size of the protein and the length of the linker used to connect the termini.

Progress in integrated assessment and modelling

Environmental processes have been modelled for decades. However. the need for integrated assessment and modeling (IAM) has,town as the extent and severity of environmental problems in the 21st Century worsens. The scale of IAM is not restricted to the global level as in climate change models, but includes local and regional models of environmental problems. This paper discusses various definitions of IAM and identifies five different types of integration that Lire needed for the effective solution of environmental problems. The future is then depicted in the form of two brief scenarios: one optimistic and one pessimistic. The current state of IAM is then briefly reviewed. The issues of complexity and validation in IAM are recognised as more complex than in traditional disciplinary approaches. Communication is identified as a central issue both internally among team members and externally with decision-makers. stakeholders and other scientists. Finally it is concluded that the process of integrated assessment and modelling is considered as important as the product for any particular project. By learning to work together and recognise the contribution of all team members and participants, it is believed that we will have a strong scientific and social basis to address the environmental problems of the 21st Century. (C) 2002 Elsevier Science Ltd. All rights reserved.

Finite element modelling of reactive fluids mixing and mineralization in pore-fluid saturated hydrothermal/sedimentary basins

We present the finite element simulations of reactive mineral carrying fluids mixing and mineralization in pore-fluid saturated hydrothermal/sedimentary basins. In particular we explore the mixing of reactive sulfide and sulfate fluids and the relevant patterns of mineralization for Load, zinc and iron minerals in the regime of temperature-gradient-driven convective flow. Since the mineralization and ore body formation may last quite a long period of time in a hydrothermal basin, it is commonly assumed that, in the geochemistry, the solutions of minerals are in an equilibrium state or near an equilibrium state. Therefore, the mineralization rate of a particular kind of mineral can be expressed as the product of the pore-fluid velocity and the equilibrium concentration of this particular kind of mineral Using the present mineralization rate of a mineral, the potential of the modern mineralization theory is illustrated by means of finite element studies related to reactive mineral-carrying fluids mixing problems in materially homogeneous and inhomogeneous porous rock basins.

Effect of shear stress on expression of a recombinant protein by Chinese hamster ovary cells

A flow chamber was used to impart a steady laminar shear stress on a recombinant Chinese hamster ovary (CHO) cell line expressing human growth hormone (hGH). The cells were subjected to shear stress ranging from 0.005 to 0.80 N m(-2). The effect of shear stress on the cell specific glucose uptake, cell specific hGH, and lactate productivity rates were calculated. No morphological changes to the cells were observed over the range of shear stresses examined. When the cells were subjected to 0.10 N m(-2) shear in protein-free media without Pluronic F-68, recombinant protein production ceased with no change in cell morphology, whereas control cultures were expressing hGH at 0.35 mug/10(6) cells/h. Upon addition of the shear protectants, Pluronic F-68 (0.2% [w/v]) or fetal bovine serum (1.0% [v/v] FBS), the productivity of the cells was restored. The effect of increasing shear stress on the cells in protein-free medium containing Pluronic F-68 was also investigated. Cell specific metabolic rates were calculated for cells under shear stress and for no-shear control cultures performed in parallel, with shear stress rates expressed as a percentage of those obtained for control cultures. Upon increasing shear from 0.005 to 0.80 N m(-2), the cell specific hGH productivity decreased from 100% at 0.005 N m(-2) to 49% at 0.80 N m(-2) relative to the no-shear control. A concurrent increase in the glucose uptake rate from 115% at 0.01 N m(-2) to 142% at 0.80 N m(-2), and decreased lactate productivity from 92% to 50%, revealed a change in the yield of products from glucose compared with the static control. It was shown that shear stress, at sublytic levels in medium containing Pluronic F-68, could decrease hGH specific productivity. (C) 2002 Wiley Periodicals, Inc.

Effect of honey types and concentration on starch gelatinization

The complex and variable composition of honey, depending on source, season and processing, means different honey samples could cause variation in the characteristics of the finished product. The objective of this study was to determine how the minor components present in honey affect starch gelatinization. A Rapid Visco Analyser was used to measure changes in viscosity when unmodified maize starch was gelatinized in a honey or model sugar solution. When honey was compared to equivalent blends of sugars, there was an increase in starch viscosity with increasing levels of addition. However, at the same level, honey gave a lower viscosity than the blends of sugars. Honeys from different sources (differing in pH and amylase activity) show a varied effect on starch gelatinization, with starch viscosity increasing with addition level for six of the honeys, but decreasing with increasing addition level for two honey samples. Varying the pH also produced variation in starch gelatinization patterns between honey types. Between pH 3.0 and 4.0, starch viscosity was similar for all four honey types studied, while above this pH there were differences between all honey types. As expected, starch viscosity decreased as the solution pH neared the optimum for honey amylase activity (pH 5.3-5.6), though it did not increase as the pH moved away from the honey amylase activity optimum. Differences between honey samples, and between honey and a model sugar mixture, in their effect on starch gelatinization was attributed to honey amylase activity and the composition and concentration of minor organic compounds present. Crown Copyright (C) 2003 Published by Elsevier Ltd. on behalf of Swiss Society of Food Science and Technology

In the eye of the storm: exploring the introduction of environmental issues in the production function in Brazilian companies

This article aims to analyse the introduction of environmental issues in the context of the production function, which has been referred to as the organisational area to lead corporate environmental management. With that purpose, the theoretical references for corporate environmental management and the necessary alterations in production function have been organised to include environmental aspects, especially in terms of product and process development, quality management, and logistics. Considering that this research field still lacks empirical evidence for Brazilian companies, four case studies were conducted using companies located in the country. The environmental management maturity level of those companies tends to follow the rate with which the environmental issue is introduced in production sub-areas, especially in the product development process. However, in most cases we found that the companies had difficulties in structuring the insertion of the environmental dimension in logistics. The final notes point out the distance observed between what is recommended by international literature and the reality of Brazilian companies in the challenge of making the production function environmentally friendly.

ON THE DIFFERENCES BETWEEN THE MARGINAL PRODUCT OF CAPITAL ACROSS COUNTRIES

We extended the standard neoclassical model of investment for the case of an open economy. Our model shows that risk premium not only creates a wedge between the marginal product of capital across countries but also reduces an economy`s savings rate. A riskier market thus presents a lower income per capita, ceteris paribus. Our empirical analysis, from 1950 to 2003, lends support to the conclusion that both risk and the correction for output price to investment ratio help to explain the differentials.

Spermatophore and Gonopore Morphology of the Southwestern-Atlantic Hermit Crab Pagurus exilis (Benedict, 1892) (Anomura, Paguridae)

Marcelo A. Scelzo, Marina Z. Fantucci, and Fernando L. Mantelatto (2010) Spermatophore and gonopore morphology of the southwestern-Atlantic hermit crab Pagurus exilis (Benedict, 1892) (Anomura, Paguridae). Zoological Studies 49(3): 421-433. The form and function of the spermatophore have been used as a complementary tool in studies of the reproductive biology and systematics of hermit crabs. In this context, we describe the spermatophore and gonopore morphology of Pagurus exilis. The spermatophores were extracted from the distal part of the vas deferens of specimens collected in Argentina and Brazil. The spermatophores were composed of 3 major regions: a main ampulla (with a sperm capsule inside and an accessory ampulla at the base), a stalk, and a pedestal. Each spermatophore had a distinct dorsolateral suture line around the ampulla, where the rupture occurs to release the sperm. The spermatophore total length was 1.5 times the main ampulla length. The main ampulla was oval and slightly flattened. A triangular accessory ampulla extended from the main ampulla base to the pedestal on 1 side, and contained no to several sperm. The stalk is short and flattened, and as wide as the main ampulla. One to 3 spermatophores were found attached to each pedestal, which was almost oblong in shape. The dimensions of the spermatophore and its component parts were directly influenced by the size of the hermit crab. Gonopores of males were covered by long pappose setae, while female gonopores bore a few short cuspidate setae. Specimens from Brazil and Argentina had the same spermatophore morphology, corroborating the previously observed absence of genetic differences between the both populations. The spermatophore morphology of this species has similarities with the broad general pattern of the Paguridae, being most similar to one of the (at least) 3 patterns of spermatophore morphology described for Pa gurus. http://zoolstud.sinica.edu.tw/Journals/49.3/421.pdf

Process development for a recombinant Chinese Hamster Ovary (CHO) cell line utilizing a metal induced and amplified metallothionein expression system

The suspension Chinese Hamster Ovary cell line, 13-10-302, utilizing the metallothionein (MT) expression system producing recombinant human growth hormone (hGH) was studied in a serum-free and cadmium-free medium at different fermentation scales and modes of operation. Initial experiments were carried out to optimize the concentration of metal addition to induce the MT promoter. Subsequently, the cultivation of the 13-10-302 cell line was scaled up from spinner flasks into bioreactors, and the cultivation duration was extended with fed-batch and perfusion strategies utilizing 180 muM zinc to induce the promoter controlling expression of recombinant hGH. It was shown that a fed-batch process could increase the maximum cell numbers twofold, from 3.3 to 6.3 x 10(6) cell/mL, over those obtained in normal batch fermentations, and this coupled with extended fermentation times resulted in a fourfold increase in final hGH titer, from 135 +/- 15 to 670 +/- 70 mg/L at a specific productivity q(hGH) value of 12 pg cell(-1)d(-1). The addition of sodium butyrate increased the specific productivity of hGH in cells to a value of approximately 48 pg cell(-1)d(-1), resulting in a final hGH titer of over a gram per liter during fed-batch runs. A BioSep acoustic cell recycler was used to retain the cells in the bioreactor during perfusion operation. It was necessary to maintain the specific feeding rates (SFR) above a value of 0.2 vvd/(10(6) cell/mL) to maintain the viability and productivity of the 13-10-302 cells; under these conditions the viable cell number increased to over 107 cell/mL and resulted in a volumetric productivity of over 120 mg(hGH) L(-1)d(-1). Process development described in this work demonstrates cultivation at various scales and sustained high levels of productivity under cadmium free condition in a CHO cell line utilizing an inducible metallothionein expression system. (C) 2004 Wiley Periodicals, Inc.

Carbamazepine oxidation catalyzed by iron and manganese porphyrins supported on aminofunctionalized matrices

This work describes the catalytic activity of manganese and iron porphyrins, Mn and Fe(TFPP)Cl, covalently immobilized on the aminofunctionalized supports montmorillonite K-10 (MontX) and silica (SilX), where X= 1 or 2 represents the length of the organic chain (""arms"") binding the metalloporphyrin to the support. These systems were characterized by UV-vis and Electronic Paramagnetic Resonance (EPR), and they were used as catalysts in the oxidation of carbamazepine (CBZ) by the oxidants iodosylbenzene (PhIO) and hydrogen peroxide. The manganese porphyrin (MnP) catalysts proved to be efficient and selective for the epoxide, the main CBZ metabolite in natural systems. MnMont1 was an excellent catalyst when PhIO was used as oxidant, even better than the same MnP in homogeneous system. Supports bearing short ""arms"" led to the best yields. Although H2O2 is an environmentally friendly oxidant, low product yields were obtained when it was employed in CBZ oxidation. Fe(TFPP)CI immobilized on aminofunctionalized supports was not an efficient catalyst, probably due to the presence of Fe(H) species in the matrix, which led to the less reactive intermediate PFe(IV)(O). (c) 2007 Elsevier B.V. All rights reserved.

An environmentally friendly triphasic catalytic system: Mn(salen) occluded in membranes based on PDMS/PVA

The commercially available Jacobsen catalyst, Mn(salen), was occluded in hybrid polymeric membranes based on poly(dimethylsiloxane) (PDMS) and poly(vinyl alcohol) (PVA). The obtained systems were characterized by UV-vis spectroscopy and SEM techniques. The membranes were used as a catalytic barrier between two different phases: an organic substrate phase (cyclooctene or styrene) in the absence of solvent, and an aqueous solution of either t-BuOOH or H(2)O(2). Membranes containing different percentages of PVA were prepared, in order to modulate their hydrophilic/hydrophobic swelling properties. The occluded complex proved to be an efficient catalyst for the oxidation of alkenes. The new triphasic system containing a cheap and easily available catalyst allowed substrate oxidation and easy product separation using ""green"" oxidants. (C) 2010 Elsevier B.V. All rights reserved.

First detection of lead in black paper from intraoral film An environmental concern

Lead (Pb) contamination in the black paper that recovers intraoral films (BKP) has been investigated. BKP samples were collected from the Radiology Clinics of the Dental School of Ribeirao Preto, University of Sao Paulo, Brazil. For sake of comparison, four different methods were used. The results revealed the presence of high lead levels, well above the maximum limit allowed by the legislation. Pb contamination levels achieved after the following treatments: paper digestion in nitric acid, microwave treatment, DIN38414-54 method and TCLP method were 997 mu g g(-1), 189 mu g g(-1), 20.8 mu g g(-1), and 54.0 mu g g(-1), respectively. Flame atomic absorption spectrometry (FAAS) and inductively coupled plasma mass spectrometry (ICP-MS) were employed for lead determination according to the protocols of the applied methods. Lead contamination in used BKP was confirmed by scanning electron microscopy coupled with energy dispersive X-ray microanalysis (SEM-EDS). All the SEM imaging was carried out in the secondary electron mode (SE) and backscattered-electron mode (QBSD) following punctual X-ray fluorescence spectra. Soil contamination derived from this product revealed the urgent need of addressing this problem. These elevated Pb levels, show that a preliminary treatment of BKP is mandatory before it is disposed into the common trash. The high lead content of this material makes its direct dumping into the environment unwise. (C) 2009 Elsevier B.V. All rights reserved.