Characterization of Staphylococcus species by SDS–PAGE of Whole-Cell and Extracellular Proteins

In this study, a total of fifteen staphylococcal strains belonging to different species were characterized by whole-cell and extracellular protein profiles using sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE). The results are presented as dendrograms after quantitative analysis of the band patterns with a computer program. Visual inspection of protein bands and cluster analysis of protein patterns of to be used 15 strains, representing 10 Staphylococcus species, showed that whole-cell and extracellular protein profiles differed in several protein bands in Staphylococcus aureus, S. epidermidis, S. simulans, and other species of Staphylococcus; however, the differences were insufficient for reliable differentiation of Staphylococcus species by the SDS–PAGE method.     

Cross-reactivity of some antibodies to human epitopes with shrimp Pandalus borealis proteins: a possible aid in validation and characterization of crustacean cells in vitro

Cell characterization of primary cultures in vertebrates is well established but not in marine invertebrates. This fact is hampering advances in the development of tissue cultures from this species. In the present study, a panel of antibodies to structural proteins, stress proteins, oncogenes and proliferation antigens, developed against mammalian antigens, were tested in paraffin sections of the crustacean Pandalus borealis tissues. Several tissues were analysed: hepatopancreas, gills, ovaries, epithelium under the cuticle and abdominal muscle. Specific antibodies to crustacean proteins are not commercially available. The immunocytochemical results show that antibodies to human epitopes cross-react with antigens in the crustacean Pandalus borealis indicating that some cellular proteins are highly conserved in evolution. Cytokeratin, proliferating cell nuclear antigen, ras and p-glycoprotein were detected by immunocytochemistry in Pandalus borealis. No immunoreactivity for Ki-67 and metallothionein was observed. This system can help in validation and characterization of invertebrate cultures.     

Application of protease from Nocardiopsis sp. as a laundry detergent additive

The application of protease as a laundry detergent additive from a newly isolated Nocardiopsis sp., isolated from a soil sample collected in Northeast Brazil is reported. The optimal pH and temperature for protease activity were pH 10.5 and 50 °C, respectively. The enzyme was stable in a long-term incubation, showed 73.5% of initial activity at pH 10.5 and 61.7% at pH 12.0 for 120 min. Approximately 60% of initial activity remained after 120 min at 50 °C or after 30 min at 80 °C. Almost 87% of enzyme activity was retained in the presence of 10% (v/v) of peroxide at 40 °C, after 1 h. The protease also was stable in the presence of oxidants and surfactants such as SDS, saponin, Tween 20 and Tween 80 after 30 min. In the presence of Omo^®, the enzyme retained 64% of its activity at 40 °C for 1 h. An increase in the proteolytic activity (6–17%) was observed with K⁺, Na⁺, and Mg⁺⁺ ions. At pH 8.0, the protease hydrolysed casein maximally (50 U/mg).     

Markovian models for the developmental study of social behavior

Behavioral development involves changes in the probabilities of both social and nonsocial activities and the sequential pattern of activities over time. A number of methods have been offered for the analysis of these patterns of behavioral sequences. However, there continue to be problematic issues, including the analysis of nonstationary data; accommodation of changes in patterns within an observation period, or over repeated observations or age; and identification of differences in pattern changes between individuals or groups, and the factors responsible for these differences. In this work, we analyze data from 15 young monkeys (Macaca nemestrina) using classification and Markovian methods, including a new approach to nonstationary data called the double-chain Markov model (DCCM). These methods allowed us to identify differences in behavior patterns that differentiate between normal subjects and those presenting developmental anomalies.     

On-line monitoring of the methanol concentration in Pichia pastoris cultures producing an heterologous lipase by sequential injection analysis

An automated sequential injection analysis (SIA) system using stop-flow technique was developed to determine methanol concentration by means of the enzymatic reactions of alcohol oxidase and peroxidase. Its application as an on-line device for monitoring Pichia pastoris fermentations producing an heterologous protein was demonstrated. Linear response, observed up to 2 g l^–1, was reached by including a dilution chamber in the SIA manifold. The sampling frequency was 7 analyses per hour with a relative standard deviation lower than 4%.     

Molecular cloning, expression, purification, and mass spectrometric characterization of 3C-like protease of SARS coronavirus

Severe acute respiratory syndrome (SARS) is an acute respiratory illness, which has broken out in China. It has been known that SARS coronavirus (SARS_CoV) is a novel human coronavirus and is responsible for SARS infection. Belonging to one of the major proteins associated with SARS_CoV, SARS 3C-like protease (SARS_3CL(pro)) functions as a cysteine protease engaging in the proteolytic cleavage of the viral precursor polyprotein to a series of functional proteins required for coronavirus replication and is considered as an appealing target for designing anti-SARS agents. To facilitate the studies regarding the functions and structures of SARS_3CL(pro), in this report the synthetic genes encoding 3CL(pro) of SARS_CoV were assembled, and the plasmid was constructed using pQE30 as vector and expressed in Escherichia coli M15 cells. The highly yielded ( approximately 15mg/L) expressed protease was purified by use of NTA-Ni(2+) affinity chromatography and FPLC system, and its sequence was determined by LC/MS with the residue coverage of 46.4%.     

Proteolytic activity of a yeast cell wall lytic Arthrobacter species

AIMS: To investigate properties of the proteolytic activity of a yeast cell wall lytic soil bacterium identified as an Arthrobacter species. METHODS AND RESULTS: The organism was grown at pH 7.5 and 30 degrees C in shake flasks on media with different complex subtrates. Highest proteolytic activity assayed with azocaseine was detected in media with wheat gluten. In addition, l-leucine, l-alanine exopeptidase activity and esterase activity were found. The proteolytic activity showed stability up to pH 12, with a maximum at pH 11. The temperature optimum was at 55 degrees C, but there was a loss in enzyme activity of 50% within 2 h. The proteolytic activity was inhibited by 3,4-dichloroisocumarin, whereas there was little or no effect with EDTA, pepstatin A or E64. CONCLUSIONS: The proteolytic activity is highly alkaline stable. The formation of the enzyme can be induced by media with high protein content.     

Advances in biotreatment of acid mine drainage and biorecovery of metals: 1. Metal precipitation for recovery and recycle

Acid mine drainage (AMD), an acidic metal-bearingwastewater, poses a severe pollution problem attributedto post mining activities. The metals usuallyencountered in AMD and considered of concern for riskassessment are arsenic, cadmium, iron, lead, manganese,zinc, copper and sulfate. The pollution generated byabandoned mining activities in the area of Butte, Montanahas resulted in the designation of the Silver Bow Creek–ButteArea as the largest Superfund (National Priorities List) sitein the U.S. This paper reports the results of bench-scalestudies conducted to develop a resource recovery basedremediation process for the clean up of the Berkeley Pit.The process utilizes selective, sequential precipitation (SSP)of metals as hydroxides and sulfides, such as copper, zinc,aluminum, iron and manganese, from the Berkeley Pit AMDfor their removal from the water in a form suitable foradditional processing into marketable precipitates and pigments.The metal biorecovery and recycle process is based on completeseparation of the biological sulfate reduction step and themetal precipitation step. Hydrogen sulfide produced in the SRBbioreactor systems is used in the precipitation step to forminsoluble metal sulfides. The average metal recoveries usingthe SSP process were as follows: aluminum (as hydroxide) 99.8%,cadmium (as sulfide) 99.7%, cobalt (as sulfide) 99.1% copper(as sulfide) 99.8%, ferrous iron (sulfide) 97.1%, manganese(as sulfide) 87.4%, nickel (as sulfide) 47.8%, and zinc (as sulfide)100%. The average precipitate purity for metals, copper sulfide,ferric hydroxide, zinc sulfide, aluminum hydroxide and manganesesulfide were: 92.4, 81.5, 97.8, 95.6 , 92.1 and 75.0%, respectively.The final produced water contained only calcium and magnesiumand both sulfate and sulfide concentrations were below usablewater limits. Water quality of this agriculturally usable watermet the EPA's gold standard criterion.     

Functional priorities in LCA and design for environment

Aim, Scope and Background  The interest in environmental questions has increased enormously during the last decade. Environmental protection has become an issue of strategic importance within the manufacturing industry and many companies are now working in the field of Design for Environment (DFE). The main purpose of DFE is to create products and services for achieving a sustainable society. Designers are widely believed to have a key role in adapting products to a sustainable society and one of the major instruments in the context of Design for Environment is Life Cycle Assessment (LCA). However, product development creates particular challenges for incorporating environmental issues that combine functional and environmental assessment. A natural and important part of product design is to define and analyse the functions of the product. Consequently, the functional unit in LCA is a core issue in DFE. Most recent research in DFE has focused on how to reduce the environmental impact of products throughout their life-cycle by addressing environmental aspects, while little attention has been given to the functionality of the product. Additionally, early product development phases, so called re-think phases, are considered to have the influence on major changes in products in general. These phases have thus the highest potential for changing products and product systems towards a sustainable development. Main Features  This paper discusses an extended functional representation in design for environment methods to evaluate sustainable design solutions, especially in early (re-think) phases of product design. Based on engineering-design science and several case studies, a concept has been developed describing how functional preferences can be visualised in design for environment and product development. In addition, the functional unit in LCA is discussed. The concept is called Functional Profile (FP) and is additionally exemplified in a case study on radio equipment. Discussion  The new functional characterisation concept helps identify functional priorities in design for environment. The Functional Profile is a structured, systematic and creative concept for identifying the necessary functions of a new product. The FP is envisioned to complement existing design for environment methods, not to replace them. Instead of being a product-development tool or method, the concept is an approach that increases understanding of inter-reactions between functional characteristics of products and their environmental characteristics, which furthermore facilitates trade-off decisions. One of the objectives behind the concept is to highlight the importance of balancing functional requirements and environmental impacts, presenting both the advantages and disadvantages of the product. Outlook  A second paper will be produced to complement the functional-environmental characterisation concept in early product development phase, presenting the environmental characterisation part and illustrating correlations between the functional and environmental sides.