A specific alkaline phosphatase from Saccharomyces cerevisiae with protein phosphatase activity

In this paper, specific PHO13 alkaline phosphatase from Saccharomyces cerevisiae was demonstrated to possess phosphoprotein phosphatase activity on the phosphoseryl proteins histone II-A and casein. The enzyme is a monomeric protein with molecular mass of 60 kDa and hydrolyzes p-nitrophenyl phosphate with maximal activity at pH 8.2 with strong dependence on Mg²⁺ ions and an apparent K_m of 3.6×10⁻⁵ M. No other substrates tested except phosphorylated histone II-A and casein were hydrolyzed at any significant rate. These data suggest that the physiological role of the p-nitrophenyl phosphate-specific phosphatase may involve participation in reversible protein phosphorylation.     

Deciphering role of technical bioprocess parameters for bioethanol production using microalgae

Microalgae biomass is considered an important feedstock for biofuels and other bioactive compounds due to its faster growth rate, high biomass production and high biomolecules accumulation over first and second-generation feedstock. This research aimed to maximize the specific growth rate of fresh water green microalgae Closteriopsis acicularis, a member of family Chlorellaceae under the effect of pH and phosphate concentration to attain enhanced biomass productivity. This study investigates the individual and cumulative effect of phosphate concentration and pH on specific growth characteristics of Closteriopsis acicularis in autotrophic mode of cultivation for bioethanol production. Central-Composite Design (CCD) strategy and Response Surface Methodology (RSM) was used for the optimization of microalga growth and ethanol production under laboratory conditions. The results showed that high specific growth rate and biomass productivity of 0.342 day⁻¹ and 0.497 g L⁻¹ day⁻¹ respectively, were achieved at high concentration of phosphate (0.115 g L⁻¹) and pH (9) at 21st day of cultivation. The elemental composition of optimized biomass has shown enhanced elemental accumulation of certain macro (C, O, P) and micronutrients (Na, Mg, Al, K, Ca and Fe) except for nitrogen and sulfur. The Fourier transform infrared spectroscopic analysis has revealed spectral peaks and high absorbance in spectral range of carbohydrates, lipids and proteins, in optimized biomass. The carbohydrates content of optimized biomass was observed as 58%, with 29.3 g L⁻¹ of fermentable sugars after acid catalyzed saccharification. The bioethanol yield was estimated as 51 % g ethanol/g glucose with maximum of 14.9 g/L of bioethanol production. In conclusion, it can be inferred that high specific growth rate and biomass productivity can be achieved by varying levels of phosphate concentration and pH during cultivation of Closteriopsis acicularis for improved yield of microbial growth, biomass and bioethanol production.     

Contents of macro-, microelements and long-lived radionuclides in the medicinal plants belonging to the wetland community of Siberian region,Russia

Studied contents of macro-, microelements and long-lived radionuclides in the following medicinal plants belonging to the wetland community of Siberian region, Russia: Sphagnum fuscum (Schimp.) Klinggr., Sphagnum balticum (Russ.) Russ. ex С. Jens., Polytrichum strictum Brid., Pinus sylvestris L., Pinus sibirica Du Tour, Betula pubescens Ehrh., Andromeda polifolia L., Ledum palustre L., Oxycoccus palustris Pers., Rubus chamaemorus L., Vaccinium uliginosum L., Comarum palustre L., Chamaedaphne calyculata (L.) Moench. Samples were collected in the sphagnum wetlands of Tomsk region and Khanty-Mansi Autonomous Okrug. For the purpose of research, we used roots, shoots and bark of the plants in the Pinophyta and Magnoliophyta species and turf of the Bryophyta species. The content of macro-, microelements in the plants was determined by an atomic emission spectrometer with inductively coupled plasma “iCAP 6300 Duo” of the “Thermo Scientific” company. The specific activities of the long-lived radionuclides Th-232, K-40, Ra-226, Cs-137 were measured by a gamma-ray spectrometric complex with a high purity germanium semiconductor detector by “ORTEC” (AMETEK) and a digital analyser “ORTEC DSPEC LF”. The obtained results of the element content are useful when selecting a medicinal plant species (or part thereof) which may be used in preparation of new medicinal drugs.     

Interactive effects of elevated CO2 and drought stress on leaf water potential and growth in Caragana intermedia

We studied the responses of leaf water potential (Ψ_w), morphology, biomass accumulation and allocation, and canopy productivity index (CPI) to the combined effects of elevated CO₂ and drought stress in Caragana intermedia seedlings. Seedlings were grown at two CO₂ concentrations (350 and 700 μmol mol⁻¹) interacted with three water regimes (60–70%, 45–55%, and 30–40% of field capacity of soil). Elevated CO₂ significantly increased Ψ_w, decreased specific leaf area (SLA) and leaf area ratio (LAR) of drought-stressed seedlings, and increased tree height, basal diameter, shoot biomass, root biomass as well as total biomass under the all the three water regimes. Growth responses to elevated CO₂ were greater in well-watered seedlings than in drought-stressed seedlings. CPI was significantly increased by elevated CO₂, and the increase in CPI became stronger as the level of drought stress increased. There were significant interactions between elevated CO₂ and drought stress on leaf water potential, basal diameter, leaf area, and biomass accumulation. Our results suggest that elevated CO₂ may enhance drought avoidance and improved water relations, thus weakening the effect of drought stress on growth of C. intermedia seedings.     

Upstream processes in antibody production: evaluation of critical parameters

The demand for monoclonal antibody for therapeutic and diagnostic applications is rising constantly which puts up a need to bring down the cost of its production. In this context it becomes a prerequisite to improve the efficiency of the existing processes used for monoclonal antibody production. This review describes various upstream processes used for monoclonal antibody production and evaluates critical parameters and efforts which are being made to enhance the efficiency of the process. The upstream technology has tremendously been upgraded from host cells used for manufacturing to bioreactors type and capacity. The host cells used range from microbial, mammalian to plant cells with mammalian cells dominating the scenario. Disposable bioreactors are being promoted for small scale production due to easy adaptation to process validation and flexibility, though they are limited by the scale of production. In this respect Wave bioreactors for suspension culture have been introduced recently. A novel bioreactor for immobilized cells is described which permits an economical and easy alternative to hollow fiber bioreactor at lab scale production. Modification of the cellular machinery to alter their metabolic characteristics has further added to robustness of cells and perks up cell specific productivity. The process parameters including feeding strategies and environmental parameters are being improved and efforts to validate them to get reproducible results are becoming a trend. Online monitoring of the process and product characterization is increasingly gaining importance. In total the advancement of upstream processes have led to the increase in volumetric productivity by 100-fold over last decade and make the monoclonal antibody production more economical and realistic option for therapeutic applications.     

DNA technological progress toward advanced diagnostic tools to support human hookworm control

Blood-feeding hookworms are parasitic nematodes of major human health importance. Currently, it is estimated that 740 million people are infected worldwide, and more than 80 million of them are severely affected clinically by hookworm disease. In spite of the health problems caused and the advances toward the development of vaccines against some hookworms, limited attention has been paid to the need for improved, practical methods of diagnosis. Accurate diagnosis and genetic characterization of hookworms is central to their effective control. While traditional diagnostic methods have considerable limitations, there has been some progress toward the development of molecular-diagnostic tools. The present article provides a brief background on hookworm disease of humans, reviews the main methods that have been used for diagnosis and describes progress in establishing polymerase chain reaction (PCR)-based methods for the specific diagnosis of hookworm infection and the genetic characterisation of the causative agents. This progress provides a foundation for the rapid development of practical, highly sensitive and specific diagnostic and analytical tools to be used in improved hookworm prevention and control programmes.     

Toward practical, DNA-based diagnostic methods for parasitic nematodes of livestock--bionomic and biotechnological implications

Parasitic nematodes of livestock have a major economic impact worldwide. In spite of the health problems caused by nematodes and advances toward the development of vaccines and new therapeutic agents against some of them, relatively limited attention has been paid to the need for improved, practical methods of diagnosis. Accurate diagnosis and genetic characterization of parasitic nematodes of livestock are central to their effective control, particularly given the current, serious problems with anthelmintic resistance in nematode populations. Traditional diagnostic techniques have considerable limitations, and there have been some advances toward the development of molecular-diagnostic tools. This article provides a brief account of the significance of parasitic nematodes (order Strongylida), reviews the techniques that have been evaluated or used for diagnosis and describes developments in polymerase chain reaction (PCR)-based methods for the specific diagnosis of nematode infection/s and the genetic characterisation of the causative agents. The advances made in recent years provide a solid foundation for the development of practical, highly sensitive and specific diagnostic tools for epidemiological investigations and for use in control programmes.     

Haplorchis taichui, Witenberg, 1930: Development of a HAT-RAPD marker for the detection of minute intestinal fluke infection

Specific primers to determine the presence of an intestinal fluke, Haplorchis taichui, were investigated using the high annealing temperature random amplified polymorphic DNA (HAT-RAPD) PCR, and 18 arbitrary primers (Operon Technologies), to generate different polymorphic DNA profiles. Thirteen kinds of parasites were used to compare fingerprints. A 256 bp HAT-RAPD marker, generated from the OPP-11 primer, was found to be H. taichui-specific, and this marker was cloned, transformed, and sequenced. From the sequence data, a pair of primers were designed with Genetyx-MAC ver.11 and indicated as: Hap-t F 5′-GGC CAA CGC AAT CGT CAT CC-3′ and Hap-t R 5′-GCG TCG GGT TTC AGA CAT GG-3′. These specific primers were tested for efficacy and specificity by amplifying them with all 13 parasites DNAs in PCR reaction. A 256 bp amplicon was generated, which was shown to have a positive result, only for H. taichui DNA. It revealed no cross-reaction with any of the other tested parasite species. The minimum DNA template, needed for detection by PCR, was 0.1 picogram (pg). The successful development of H. taichui-specific primers is expected to be beneficial for epidemiological studies and for prevention and control of these parasitic infections.     

Use of NaCl prevents aggregation of recombinant COMP–Angiopoietin-1 in Chinese hamster ovary cells

To investigate the effect of hyperosmotic medium on production and aggregation of the variant of Angiopoietin-1 (Ang1), cartilage oligomeric matrix protein (COMP)–Ang1, in recombinant Chinese hamster ovary (CHO) cells, CHO cells were cultivated in shaking flasks. NaCl and/or sorbitol were used to raise medium osmolality in the range of 300–450 mOsm/kg. The specific productivity of COMP–Ang1, q_COMP–Ang1, increased as medium osmolality increased. At NaCl-450 mOsm/kg, the q_COMP–Ang1 was 7.7-fold higher than that at NaCl-300 mOsm/kg, while, at sorbitol-450 mOsm/kg, it was 2.9-fold higher than that at sorbitol-300 mOsm/kg. This can be attributed to the increased relative mRNA level of COMP–Ang1 at NaCl-450 mOsm/kg which was approximately 2.4-fold higher than that at sorbitol-450 mOsm/kg. Western blot analysis showed that COMP–Ang1 aggregates started to occur in the late-exponential phase of cell growth. When sorbitol was used to raise the medium osmolality, a severe aggregation of COMP–Ang1 was observed. On the other hand, when NaCl was used, the aggregation of COMP–Ang1 was drastically reduced at NaCl-400 mOsm/kg. At NaCl-450 mOsm/kg, the aggregation of COMP–Ang1 was hardly observed. This suggests that environmental conditions are critical for the aggregation of COMP–Ang1. Taken together, the use of NaCl-induced hyperosmotic medium to cell culture process turns out to be an efficient strategy for enhancing COMP–Ang1 production and reducing COMP–Ang1 aggregation.     

Specific dynamic action: a review of the postprandial metabolic response

For more than 200 years, the metabolic response that accompanies meal digestion has been characterized, theorized, and experimentally studied. Historically labeled “specific dynamic action” or “SDA”, this physiological phenomenon represents the energy expended on all activities of the body incidental to the ingestion, digestion, absorption, and assimilation of a meal. Specific dynamic action or a component of postprandial metabolism has been quantified for more than 250 invertebrate and vertebrate species. Characteristic among all of these species is a rapid postprandial increase in metabolic rate that upon peaking returns more slowly to prefeeding levels. The average maximum increase in metabolic rate stemming from digestion ranges from a modest 25% for humans to 136% for fishes, and to an impressive 687% for snakes. The type, size, composition, and temperature of the meal, as well as body size, body composition, and several environmental factors (e.g., ambient temperature and gas concentration) can each significantly impact the magnitude and duration of the SDA response. Meals that are large, intact or possess a tough exoskeleton require more digestive effort and thus generate a larger SDA than small, fragmented, or soft-bodied meals. Differences in the individual effort of preabsorptive (e.g., swallowing, gastric breakdown, and intestinal transport) and postabsorptive (e.g., catabolism and synthesis) events underlie much of the variation in SDA. Specific dynamic action is an integral part of an organism’s energy budget, exemplified by accounting for 19–43% of the daily energy expenditure of free-ranging snakes. There are innumerable opportunities for research in SDA including coverage of unexplored taxa, investigating the underlying sources, determinants, and the central control of postprandial metabolism, and examining the integration of SDA across other physiological systems.