ISRN Biochemistry The latest articles from Hindawi Publishing Corporation © 2014 , Hindawi Publishing Corporation . All rights reserved. Analysis of Mathematical Modelling on Potentiometric Biosensors Wed, 07 May 2014 12:22:24 +0000 A mathematical model of potentiometric enzyme electrodes for a nonsteady condition has been developed. The model is based on the system of two coupled nonlinear time-dependent reaction diffusion equations for Michaelis-Menten formalism that describes the concentrations of substrate and product within the enzymatic layer. Analytical expressions for the concentration of substrate and product and the corresponding flux response have been derived for all values of parameters using the new homotopy perturbation method. Furthermore, the complex inversion formula is employed in this work to solve the boundary value problem. The analytical solutions obtained allow a full description of the response curves for only two kinetic parameters (unsaturation/saturation parameter and reaction/diffusion parameter). Theoretical descriptions are given for the two limiting cases (zero and first order kinetics) and relatively simple approaches for general cases are presented. All the analytical results are compared with simulation results using Scilab/Matlab program. The numerical results agree with the appropriate theories. N. Mehala and L. Rajendran Copyright © 2014 N. Mehala and L. Rajendran. All rights reserved. Tumor Microenvironment: A New Treatment Target for Cancer Sun, 13 Apr 2014 00:00:00 +0000 Recent advances in cancer therapy encounter a bottleneck. Relapsing/recurrent disease almost always developed eventually with resistance to the initially effective drugs. Tumor microenvironment has been gradually recognized as a key contributor for cancer progression, epithelial-mesenchymal transition of the cancer cells, angiogenesis, cancer metastasis, and development of drug resistance, while dysregulated immune responses and interactions between various components in the microenvironment all play important roles. Future development of anticancer treatment should take tumor microenvironment into consideration. Besides, we also discuss the limitations of current pre-clinical testing models that mainly come from the impossibility in simulating all detailed carcinogenic mechanisms in human, especially failure to create the same tumor microenvironment. With the cumulating knowledge about tumor microenvironment, the design of a novel anticancer therapy may be facilitated and may have better chance for success in cancer eradication. Ming-Ju Tsai, Wei-An Chang, Ming-Shyan Huang, and Po-Lin Kuo Copyright © 2014 Ming-Ju Tsai et al. All rights reserved. New Lipase for Biodiesel Production: Partial Purification and Characterization of LipSB 25-4 Mon, 10 Mar 2014 09:11:45 +0000 The lipolytic activities of 300 Streptomyces isolates were determined in Tributyrin and Rhodamine-B Agar. Lipase activities were also measured with p-nitrophenyl palmitate (p-NPP) as a substrate. The strain of Streptomyces bambergiensis OC 25-4 used in this study was selected among 300 strains of Streptomyces from MUCC as the best lipase producer. The incubation conditions were optimized and the inoculum amount, incubation period, effect of carbon and nitrogen sources, and rates of MgSO4 and CaCO3 were investigated. LipSB 25-4 (the lipase produced by S. bambergiensis OC 25-4 strain) was partially purified with ammonium sulphate precipitation, dialysis, and gel filtration chromatography 2.73-fold and with 92.12 U/mg specific activity. The optimal pH and temperature for LipSB 25-4 were determined as 8.0 and 50°C, respectively. The lipase has high stability in all pH and temperature values used in this study. While LipSB 25-4 was slightly activated in the presence of β-mercaptoethanol, it was slightly reduced by PMSF. The enzyme conserved approximately 75% of its activity at the end of 60 h, in the presence of methanol and ethanol. Since LipSB 25-4 displays high activity in the thermophilic conditions and stability in the presence of organic solvents, this lipase can catalyse the biodiesel production from olive oil by the transesterification reactions. Aysel Ugur, Nurdan Sarac, Rukiye Boran, Berk Ayaz, Ozgur Ceylan, and Gulten Okmen Copyright © 2014 Aysel Ugur et al. All rights reserved. Synthetic Melatoninergic Ligands: Achievements and Prospects Sun, 23 Feb 2014 15:35:13 +0000 Pineal hormone melatonin is widely used in the treatment of disorders of circadian rhythms. The presence of melatonin receptors in various animal tissues motivates the use of this hormone in some other diseases. For this reason, in recent years investigators continued the search for synthetic analogues of melatonin which are metabolically stable and selective to receptors. This review includes recent information about the most famous melatonin analogues, their structure, properties, and physiological features of the interaction with melatonin receptors. N. V. Kostiuk, M. B. Belyakova, D. V. Leshchenko, V. V. Zhigulina, and M. V. Miniaev Copyright © 2014 N. V. Kostiuk et al. All rights reserved. Kinetic Characterization and Effect of Immobilized Thermostable β-Glucosidase in Alginate Gel Beads on Sugarcane Juice Thu, 20 Feb 2014 14:16:14 +0000 A thermostable β-glucosidase was effectively immobilized on alginate by the method of gel entrapment. After optimization of immobilized conditions, recovered enzyme activity was 60%. Optimum pH, temperature, kinetic parameters, thermal and pH stability, reusability, and storage stability were investigated. The and for immobilized β-glucosidase were estimated to be 5.0 mM and 0.64 U/ml, respectively. When comparing, free and immobilized enzyme, change was observed in optimum pH and temperature from 5.0 to 6.0 and 60°C to 80°C, respectively. Immobilized enzyme showed an increase in pH stability over the studied pH range (3.0–10.0) and stability at temperature up to 80°C. The storage stability and reusability of the immobilized β-glucosidase were improved significantly, with 12.09% activity retention at 30°C after being stored for 25 d and 17.85% residual activity after being repeatedly used for 4 times. The effect of both free and immobilized β-glucosidase enzyme on physicochemical properties of sugarcane juice was also analyzed. Keerti, Anuradha Gupta, Vinod Kumar, Ashutosh Dubey, and A. K. Verma Copyright © 2014 Keerti et al. All rights reserved. Production of Alkaline Protease by Solvent-Tolerant Alkaliphilic Bacillus circulans MTCC 7942 Isolated from Hydrocarbon Contaminated Habitat: Process Parameters Optimization Thu, 28 Nov 2013 10:36:33 +0000 In the present investigation, a newly isolated organic solvent-tolerant and alkaliphilic bacterial strain was reported from a hydrocarbon (gasoline and diesel) contaminated soil collected from the petrol station, Shirpur (India). The strain was identified as Bacillus circulans MTCC 7942, based on phenotype, biochemical, and phylogenetic analysis of 16S rRNA gene sequence. The capability of Bacillus circulans to secrete an extracellular, thermostable, alkaline protease and grow in the presence of organic solvents was explored. Bacillus circulans produced maximum alkaline protease (412 U/mL) in optimized medium (g/L): soybean meal, 15; starch, 10; KH2PO4, 1; MgSO4·7H2O, 0.05; CaCl2, 1; Na2CO3, 8; pH 10.0 at 37°C and 100 rpm. The competence of strain to grow in various organic solvents—n-octane, dodecane, n-decane, N,N-dimethylformamide, n-hexane, and dimethyl sulfoxide, establishes its potential as solvent-stable protease source for the possible applications in nonaqueous reactions and fine chemical synthesis. Ulhas Patil and Ambalal Chaudhari Copyright © 2013 Ulhas Patil and Ambalal Chaudhari. All rights reserved. Variable-Temperature Size Exclusion Chromatography for the Study of the Structural Changes in G-Quadruplex Sun, 10 Nov 2013 14:47:42 +0000 The conformational equilibria of a guanine-rich sequence found at the promoter region of the human c-kit oncogene are studied by means of circular dichroism spectroscopy (CD) and variable-temperature size exclusion chromatography (SEC). It is shown that the wild sequence ckit21 exists as a mixture of monomeric and multimeric G-quadruplexes. Appropriate mutation of several bases in the wild sequence produces the shift from parallel to antiparallel G-quadruplex, as well as the disappearance of multimeric species. The shift from the antiparallel to the parallel conformation induced by temperature is reflected in both CD and SEC profiles. Sanae Benabou, Ramon Eritja, and Raimundo Gargallo Copyright © 2013 Sanae Benabou et al. All rights reserved. Serum Prolidase Activity, Oxidant and Antioxidant Status in Nonulcer Dyspepsia and Healthy Volunteers Tue, 29 Oct 2013 13:18:01 +0000 Helicobacter pylori (H. pylori) infection is associated with increased oxidative stress and serum prolidase activity (SPA) in many diseases. We aimed to observe SPA and oxidative stress in nonulcer dyspepsia (NUD) infected with and without H. pylori among eastern Indians. 106 patients with H. pylori positive NUD, 82 patients with H. pylori negative NUD, and 50 healthy individuals were selected. SPA, total antioxidant capacity (TAOC), and total oxidant status (TOS) were measured with the use of spectrophotometer and an automated measurement method. SPA, TOS, and oxidative stress index (OSI) were significantly higher in patients with H. pylori positive than H. pylori negative NUD and healthy individuals (all ), whereas TAOC was significantly lower (). Nonsignificant, increased SPA ( value = 0.6083) and decreased TAOC ( value = 0.1186) were observed in patients with H. pylori negative NUD than healthy individuals, while increased TOS and OSI were significant (). Weak, nonsignificant correlations were observed between serum prolidase activity and TAOC, TOS, and OSI in H. pylori positive cases. Thus, increased SPA along with increased oxidative stress was observed, which seem to be closely associated with H. pylori infection. SPA and oxidative stress seem to be used as biomarkers for H. pylori infection in NUD. Shweta Kumari, Akhilesh Kumar Verma, Sumit Rungta, Rahul Mitra, Ragini Srivastava, and Narender Kumar Copyright © 2013 Shweta Kumari et al. All rights reserved. Evaluation of the In Vitro Efficacy of Artemisia annua, Rumex abyssinicus, and Catha edulis Forsk Extracts in Cancer and Trypanosoma brucei Cells Sun, 22 Sep 2013 15:39:23 +0000 The current drugs against sleeping sickness are derived from cancer chemotherapeutic approaches. Herein, we aimed at evaluating the in vitro effect of alcoholic extracts of Artemisia annua (AMR), Rumex abyssinicus (RMA), and Catha edulis Forsk (CEF) on proliferation/viability of 1321N1 astrocytoma, MCF-7 breast cancer, THP-1 leukemia, and LNCaP, Du-145, and PC-3 prostate cancer cells and on Trypanosoma brucei cells. Proliferation of tumor cells was evaluated by WST-1 assay and viability/behaviour of T. brucei by cell counting and light microscopy. CEF was the most efficient growth inhibitor in comparison to AMR and RMA. Nevertheless, in LNCaP and THP-1 cells, all extracts significantly inhibited tumor growth at 3 μg/mL. All extracts inhibited proliferation of T. brucei cells in a concentration-dependent manner. Microscopic analysis revealed that 95% of the T. brucei cells died when exposed to 33 μg/mL CEF for 3 hrs. Similar results were obtained using 33 μg/mL AMR for 6 hrs. In case of RMA, however, higher concentrations were necessary to obtain similar effects on T. brucei. This demonstrates the antitumor efficacy of these extracts as well as their ability to dampen viability and proliferation of T. brucei, suggesting a common mechanism of action on highly proliferative cells, most probably by targeting cell metabolism. Netsanet Worku, Andualem Mossie, August Stich, Arwid Daugschies, Susanne Trettner, Nasr Y. A. Hemdan, and Gerd Birkenmeier Copyright © 2013 Netsanet Worku et al. All rights reserved. Human Sprouty1 Suppresses Urokinase Receptor-Stimulated Cell Migration and Invasion Thu, 12 Sep 2013 12:18:53 +0000 The urokinase-type plasminogen activator receptor (uPAR) has been implicated in several processes in tumor progression including cell migration and invasion in addition to initiation of signal transduction. Since uPAR lacks a transmembrane domain, it uses the interaction with other proteins to modulate intracellular signal transduction. We have previously identified hSpry1 as a partner protein of uPAR, suggesting a physiological role for hSpry1 in the regulation of uPAR signal transduction. In this study, hSpry1 was found to colocalize with uPAR upon stimulation with epidermal growth factor (EGF), urokinase (uPA), or its amino terminal fragment (uPA-ATF), implicating a physiological role of hSpry1 in regulation of uPAR signalling pathway. Moreover, hSpry1 was able to inhibit uPAR-stimulated cell migration in HEK293/uPAR, breast carcinoma, and colorectal carcinoma cells. In addition, hSpry1 was found to inhibit uPAR-stimulated cell invasion in breast carcinoma and osteosarcoma cell lines. Increasing our understanding of how hSpry1 negatively regulates uPAR-stimulated cellular functions may determine a distinctive role for hSpry1 in tumour suppression. Ahmed H. Mekkawy and David L. Morris Copyright © 2013 Ahmed H. Mekkawy and David L. Morris. All rights reserved. A Sequence-Specific Nicking Endonuclease from Streptomyces: Purification, Physical and Catalytic Properties Wed, 21 Aug 2013 09:59:07 +0000 A sequence-specific nicking endonuclease from Streptomyces designated as DC13 was purified to near homogeneity. Starting with 30 grams of wet cells, the enzyme was purified by ammonium sulfate fractionation, DEAE cellulose, and phenyl-Sepharose chromatography. The purified protein had a specific activity 1000 units/mg and migrated on SDS-PAGE gel with an estimated molecular weight of 71 kDa. Determination of subunit composition by gel filtration chromatography indicated that the native enzyme is a monomer. When incubated with different DNA substrates including pBluescript II KS, pUC118, pET-15b, and pET-26b, the enzyme converted these supercoiled plasmids to a mixture of open circular and linear DNA products, with the open circular DNA as the major cleavage product. Analysis of the kinetic of DNA cleavage showed that the enzyme appeared to cleave super-coiled plasmid in two distinct steps: a rapid cleavage of super-coiled plasmid to an open circular DNA followed a much slower step to linear DNA. The DNA cleavage reaction of the enzyme required Mg2+ as a cofactor. Based on the monomeric nature of the enzyme, the kinetics of DNA cleavage exhibited by the enzyme, and cofactor requirement, it is suggested here that the purified enzyme is a sequence-specific nicking endonuclease that is similar to type IIS restriction endonuclease. Peechapack Somyoonsap, Vichein Kitpreechavanich, and Somchai Pornbanlualap Copyright © 2013 Peechapack Somyoonsap et al. All rights reserved. Biochemical Studies on Methylglyoxal-Mediated Glycated Histones: Implications for Presence of Serum Antibodies against the Glycated Histones in Patients with Type 1 Diabetes Mellitus Wed, 07 Aug 2013 13:08:21 +0000 Reactive carbonyl species (RCS) mainly reacts with lysine and arginine residues of proteins to form advanced glycation end products (AGEs). Histone was glycoxidated with glyoxal and methylglyoxal. It was characterized by polyacrylamide gel electrophoresis and quenching studies involving penicillamine and aminoguanidine as carbonyl scavengers. Further characterization of histone modified with methylglyoxal was done by UV, fluorescence, and IR spectrophotometry. Spectral analysis of the protein clearly demonstrates structural perturbation in the histone by methylglyoxal. Methylglyoxal-induces cross-linking in the protein leading to aggregation. Role of methylglyoxal mediated glycoxidation of histone in type 1 diabetes was also undertaken. Antibodies were detected against glycoxidated histone in sera of type 1 diabetes patients by solid-phase enzyme immunoassay. The findings indicate that as a result of structural perturbation in histone by methylglyoxal, the modified histone may be involved in production of serum antibodies in the diabetes patients. Nadeem A. Ansari and Debabrata Dash Copyright © 2013 Nadeem A. Ansari and Debabrata Dash. All rights reserved. Cellular and Biochemical Mechanisms of the Retroviral Restriction Factor SAMHD1 Wed, 17 Jul 2013 08:21:17 +0000 Replication of HIV-1 and other retroviruses is dependent on numerous host proteins in the cells. Some of the host proteins, however, function as restriction factors to block retroviral infection of target cells. The host protein SAMHD1 has been identified as the first mammalian deoxynucleoside triphosphate triphosphohydrolase (dNTPase), which blocks the infection of HIV-1 and other retroviruses in non-cycling immune cells. SAMHD1 protein is highly expressed in human myeloid-lineage cells and CD4+ T-lymphocytes, but its retroviral restriction function is only observed in noncycling cells. Recent studies have revealed biochemical mechanisms of SAMHD1-mediated retroviral restriction. In this review, the latest progress on SAMHD1 research is summarized and the mechanisms by which SAMHD1 mediates retroviral restriction are analyzed. Although the physiological function of SAMHD1 is largely unknown, this review provides perspectives about the role of endogenous SAMHD1 protein in maintaining normal cellular function, such as nucleic acid metabolism and the proliferation of cells. Li Wu Copyright © 2013 Li Wu. All rights reserved. Nanoparticles for Brain Drug Delivery Tue, 21 May 2013 19:05:54 +0000 The central nervous system, one of the most delicate microenvironments of the body, is protected by the blood-brain barrier (BBB) regulating its homeostasis. BBB is a highly complex structure that tightly regulates the movement of ions of a limited number of small molecules and of an even more restricted number of macromolecules from the blood to the brain, protecting it from injuries and diseases. However, the BBB also significantly precludes the delivery of drugs to the brain, thus, preventing the therapy of a number of neurological disorders. As a consequence, several strategies are currently being sought after to enhance the delivery of drugs across the BBB. Within this review, the recently born strategy of brain drug delivery based on the use of nanoparticles, multifunctional drug delivery systems with size in the order of one-billionth of meters, is described. The review also includes a brief description of the structural and physiological features of the barrier and of the most utilized nanoparticles for medical use. Finally, the potential neurotoxicity of nanoparticles is discussed, and future technological approaches are described. The strong efforts to allow the translation from preclinical to concrete clinical applications are worth the economic investments. Massimo Masserini Copyright © 2013 Massimo Masserini. All rights reserved. 1.2, Cell Proliferation, and New Target in Atherosclerosis Sun, 12 May 2013 13:45:51 +0000 1.2 calcium channels are the principal proteins involved in electrical, mechanical, and/or signaling functions of the cell. 1.2 couples membrane depolarization to the transient increase in intracellular Ca2+ concentration that is a trigger for muscle contraction and CREB-dependent transcriptional activation. The CACNA1C gene coding for the 1.2 pore-forming subunit is subject to extensive alternative splicing. This review is the first attempt to follow the association between cell proliferation, 1.2 expression and splice variation, and atherosclerosis. Based on insights into the association between the atherosclerosis-induced molecular remodeling of 1.2, proliferation of vascular smooth muscle cells, and CREB-dependent transcriptional signaling, this review will give a perspective outlook for the use of the CACNA1C exon skipping as a new potential gene therapy approach to atherosclerosis. Nikolai M. Soldatov Copyright © 2013 Nikolai M. Soldatov. All rights reserved. Transsulfuration Is a Significant Source of Sulfur for Glutathione Production in Human Mammary Epithelial Cells Wed, 06 Mar 2013 13:20:48 +0000 The transsulfuration pathway, through which homocysteine from the methionine cycle provides sulfur for cystathionine formation, which may subsequently be used for glutathione synthesis, has not heretofore been identified as active in mammary cells. Primary human mammary epithelial cells (HMEC’s) were labeled with -methionine for 24 hours following pretreatment with a vehicle control, the cysteine biosynthesis inhibitor propargylglycine or the gamma-glutamylcysteine synthesis inhibitor buthionine sulfoximine. Cell lysates were prepared and reacted with glutathione-S-transferase and the fluorescent labeling compound monochlorobimane to form a fluorescent glutathione-bimane conjugate. Comparison of fluorographic and autoradiographic images indicated that glutathione had incorporated -methionine demonstrating that functional transsulfuration occurs in mammary cells. Pathway inhibitors reduced incorporation by roughly 80%. Measurement of glutathione production in HMEC’s treated with and without hydrogen peroxide and/or pathway inhibitors indicates that the transsulfuration pathway plays a significant role in providing cysteine for glutathione production both normally and under conditions of oxidant stress. Andrea D. Belalcázar, John G. Ball, Leslie M. Frost, Monica A. Valentovic, and John Wilkinson IV Copyright © 2013 Andrea D. Belalcázar et al. All rights reserved. Metabolic Regulation of a Bacterial Cell System with Emphasis on Escherichia coli Metabolism Mon, 18 Feb 2013 14:34:34 +0000 It is quite important to understand the overall metabolic regulation mechanism of bacterial cells such as Escherichia coli from both science (such as biochemistry) and engineering (such as metabolic engineering) points of view. Here, an attempt was made to clarify the overall metabolic regulation mechanism by focusing on the roles of global regulators which detect the culture or growth condition and manipulate a set of metabolic pathways by modulating the related gene expressions. For this, it was considered how the cell responds to a variety of culture environments such as carbon (catabolite regulation), nitrogen, and phosphate limitations, as well as the effects of oxygen level, pH (acid shock), temperature (heat shock), and nutrient starvation. Kazuyuki Shimizu Copyright © 2013 Kazuyuki Shimizu. All rights reserved. Role of Elicitors in Inducing Resistance in Plants against Pathogen Infection: A Review Mon, 28 Jan 2013 08:42:21 +0000 Disease control is largely based on the use of fungicides, bactericides, and insecticides—chemical compounds toxic to plant invaders, causative agents, or vectors of plant diseases. However, the hazardous effect of these chemicals or their degradation products on the environment and human health strongly necessitates the search for new, harmless means of disease control. There must be some natural phenomenon of induced resistance to protect plants from disease. Elicitors are compounds, which activate chemical defense in plants. Various biosynthetic pathways are activated in treated plants depending on the compound used. Commonly tested chemical elicitors are salicylic acid, methyl salicylate, benzothiadiazole, benzoic acid, chitosan, and so forth which affect production of phenolic compounds and activation of various defense-related enzymes in plants. Their introduction into agricultural practice could minimize the scope of chemical control, thus contributing to the development of sustainable agriculture. This paper chiefly highlights the uses of elicitors aiming to draw sufficient attention of researchers to the frontier research needed in this context. Meenakshi Thakur and Baldev Singh Sohal Copyright © 2013 Meenakshi Thakur and Baldev Singh Sohal. All rights reserved. Locus-Specific Biochemical Epigenetics/Chromatin Biochemistry by Insertional Chromatin Immunoprecipitation Thu, 10 Jan 2013 14:29:02 +0000 Comprehensive understanding of regulation mechanisms of biological phenomena mediated by functions of genomic DNA requires identification of molecules bound to genomic regions of interest in vivo. However, nonbiased methods to identify molecules bound to specific genomic loci in vivo are limited. To perform biochemical and molecular biological analysis of specific genomic regions, we developed the insertional chromatin immunoprecipitation (iChIP) technology to purify the genomic regions of interest. We applied iChIP to direct identification of components of insulator complexes, which function as boundaries of chromatin domain, showing that it is feasible to directly identify proteins and RNA bound to a specific genomic region in vivo by using iChIP. In addition, recently, we succeeded in identifying proteins and genomic regions interacting with a single copy endogenous locus. In this paper, we will discuss the application of iChIP to epigenetics and chromatin research. Toshitsugu Fujita and Hodaka Fujii Copyright © 2013 Toshitsugu Fujita and Hodaka Fujii. All rights reserved. Ganglioside Biochemistry Wed, 19 Dec 2012 14:35:09 +0000 Gangliosides are sialic acid-containing glycosphingolipids. They occur especially on the cellular surfaces of neuronal cells, where they form a complex pattern, but are also found in many other cell types. The paper provides a general overview on their structures, occurrence, and metabolism. Key functional, biochemical, and pathobiochemical aspects are summarized. Thomas Kolter Copyright © 2012 Thomas Kolter. All rights reserved. Neuronal Nicotinic Receptors in Sleep-Related Epilepsy: Studies in Integrative Biology Sun, 09 Dec 2012 16:59:40 +0000 Although Mendelian diseases are rare, when considered one by one, overall they constitute a significant social burden. Besides the medical aspects, they propose us one of the most general biological problems. Given the simplest physiological perturbation of an organism, that is, a single gene mutation, how do its effects percolate through the hierarchical biological levels to determine the pathogenesis? And how robust is the physiological system to this perturbation? To solve these problems, the study of genetic epilepsies caused by mutant ion channels presents special advantages, as it can exploit the full range of modern experimental methods. These allow to extend the functional analysis from single channels to whole brains. An instructive example is autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE), which can be caused by mutations in neuronal nicotinic acetylcholine receptors. In vitro, such mutations often produce hyperfunctional receptors, at least in heterozygous condition. However, understanding how this leads to sleep-related frontal epilepsy is all but straightforward. Several available animal models are helping us to determine the effects of ADNFLE mutations on the mammalian brain. Because of the complexity of the cholinergic regulation in both developing and mature brains, several pathogenic mechanisms are possible, which also present different therapeutic implications. Andrea Becchetti Copyright © 2012 Andrea Becchetti. All rights reserved. Feeding of Bait to Snail Lymnaea acuminata and Their Effect on Certain Enzyme in the Nervous Tissue Tue, 04 Dec 2012 11:23:11 +0000 Fascioliasis, a snail-borne parasitic zoonosis, has been recognized for a long time because of its major veterinary and human impact. Different Bait formulations were fed to the snail Lymnaea acuminata in clear glass aquaria having diameter of 30 cm. Snail attractant containing bait formulations was prepared from different binary combination (1 : 1 ratio) of carbohydrates (glucose, starch 10 mM) and amino acid (methionine, histidine 10 mM) in 100 ml of 2% agar solution + sublethal (20% and 60% of 24 h and 96 h LC50) doses of different molluscicides (eugenol, ferulic acid, umbelliferone, and limonene). Snails fed on bait containing sub-lethal concentration of different molluscicides and the snail attractant, causing a significant inhibition in alkaline phosphatase (ALP) and acetylcholinesterase (AChE) activity in the nervous tissue of the vector snail L. acuminata. Maximum inhibition in ALP (20% of control) and AChE (49.49% of control) activity was observed in the nervous tissue of the L. acuminata exposed to 60% of 96 h LC50 of eugenol in the bait pellets containing starch + histidine, starch + methionine, respectively. Pradeep Kumar, V. K. Singh, and D. K. Singh Copyright © 2012 Pradeep Kumar et al. All rights reserved. Calpain Dysregulation in Alzheimer’s Disease Tue, 16 Oct 2012 17:35:15 +0000 Alzheimer’s disease (AD) is characterized by the presence of senile plaques and neurofibrillary tangles in the neocortex and hippocampus of AD patients. In addition, a marked decrease in synaptic contacts has been detected in these affected brain areas. Due to its prevalence in the aging population, this disease has been the focus of numerous studies. The data obtained from those studies suggest that the mechanisms leading to the formation of the hallmark lesions of AD might be linked. One of such mechanisms seems to be the dysregulation of calcium homeostasis that results in the abnormal activation of calpains. Calpains are a family of Ca2+-dependent cysteine proteases that play a key role in multiple cell functions including cell development, differentiation and proliferation, axonal guidance, growth cone motility, and cell death, among others. In this paper, we briefly reviewed data on the structure of these proteases and their regulation under normal conditions. We also summarized data underscoring the participation of calpains in the neurodegenerative mechanisms associated with AD. Adriana Ferreira Copyright © 2012 Adriana Ferreira. All rights reserved. Characterization of LlaKI, a New Metal Ion-Independent Restriction Endonuclease from Lactococcus lactis KLDS4 Sun, 30 Sep 2012 09:20:05 +0000 Requirement of divalent cations for DNA cleavage is a general feature of type II restriction enzymes with the exception of few members of this group. A new type II restriction endonuclease has been partially purified from Lactococcus lactis KLDS4. The enzyme was denoted as LlaKI and showed to recognize and cleave the same site as FokI. The enzyme displayed a denatured molecular weight of 50 kDa and behaved as a dimer in solution as evidenced by the size exclusion chromatography. To investigate the role of divalent cations in DNA cleavage by LlaKI, digestion reactions were carried out at different Mg2+, Mn2+, and Ca2+ concentrations. Unlike most of type II restriction endonucleases, LlaKI did not require divalent metal ions to cleave DNA and is one of the few metal-independent restriction endonucleases found in bacteria. The enzyme showed near-maximal levels of activity in 10 mM Tris-HCl pH 7.9, 50 mM NaCl, 10 mM MgCl2, and 1 mM dithiothreitol at 30°C. The presence of DNA modification was also determined and was correlated with the correspondent restriction enzyme. Abdelkarim Belkebir and Houssine Azeddoug Copyright © 2012 Abdelkarim Belkebir and Houssine Azeddoug. All rights reserved. Constrained Peptides as Miniature Protein Structures Wed, 26 Sep 2012 15:41:37 +0000 This paper discusses the recent developments of protein engineering using both covalent and noncovalent bonds to constrain peptides, forcing them into designed protein secondary structures. These constrained peptides subsequently can be used as peptidomimetics for biological functions such as regulations of protein-protein interactions. Hang Yin Copyright © 2012 Hang Yin. All rights reserved. Withaferin A Induces Proteasome-Dependent Degradation of Breast Cancer Susceptibility Gene 1 and Heat Shock Factor 1 Proteins in Breast Cancer Cells Sun, 02 Sep 2012 14:52:32 +0000 The purpose of this study was to examine the regulation of prosurvival factors heat shock factor 1 (HSF1) and breast cancer susceptibility gene 1 (BRCA1) by a natural withanolide withaferin A (WA) in triple negative breast cancer cell lines MDA-MB-231 and BT20. Western analysis was used to examine alternations in HSF1 and BRCA1 protein levels following WA treatment. A protein synthesis inhibitor cycloheximide and a proteasome inhibitor MG132 were used to investigate the mechanisms of HSF1 and BRCA1 regulation by WA. It was found that WA induced a dose-dependent decrease in HSF1 and BRCA1 protein levels. Further analysis showed that levels of HSF1 and BRCA1 proteins decreased rapidly after WA treatment, and this was attributed to WA-induced denaturation of HSF1 and BRCA1 proteins and subsequent degradation via proteasome-dependent, and protein-synthesis dependent mechanism. In summary, WA induces denaturation and proteasomal degradation of HSF1 and BRCA1 proteins. Further studies are warranted to examine the contribution of HSF1 and BRCA1 depletion to the anticancer effects of WA in breast cancer. Xuan Zhang, Barbara Timmermann, Abbas K. Samadi, and Mark S. Cohen Copyright © 2012 Xuan Zhang et al. All rights reserved. The Effect of the Combined Action of Roscovitine and Paclitaxel on the Apoptotic and Cell Cycle Regulatory Mechanisms in Colon and Anaplastic Thyroid Cancer Cells Thu, 30 Aug 2012 11:32:49 +0000 Aim. To study the significance of cyclin-dependent kinases (Cdks) in paclitaxel-dependent apoptosis in colon and undifferentiated thyroid cancer cells. Materials and Methods. Experiments were performed on undifferentiated thyroid carcinoma (KTC-2) and colon carcinoma (ARO) cell lines. Cells were treated with paclitaxel (Ptx) and inhibitor of Cdk, roscovitine. Cell survival test and Western blotting were used for characterization of the effects of paclitaxel and roscovitine on cancer cells. Results. It was shown that not c-Jun N-terminal kinase, but cyclin-dependent kinases are responsible for antiapoptotic Bcl-2 phosphorylation. Cdk inhibition enhanced the cytotoxic effects of Ptx at low drug concentrations. There was antagonism between Ptx and roscovitine at higher (25 nM) paclitaxel concentrations. Conclusion. Using of paclitaxel at low (2.5 to 5 nM) concentrations and roscovitine is a promising combination for further preclinical trials for the development of new therapeutic approaches to the treatment of colon and anaplastic thyroid cancer. V. V. Pushkarev, O. I. Kovzun, V. M. Pushkarev, and M. D. Tronko Copyright © 2012 V. V. Pushkarev et al. All rights reserved. The Essence of ATP Coupling Thu, 10 May 2012 08:42:10 +0000 The traditional explanation of ATP coupling is based on the raising of the equilibrium constants of the biochemical reactions. But in the frames of the detailed balance, no coupling occurs under thermodynamic equilibrium. The role of ATP in coupling is not that it provides an increase in the equilibrium constants of thermodynamically unfavorable reactions but that the unfavorable reactions are replaced by other reactions which kinetically are more favorable and give rise to the same products. The coupling with ATP hydrolysis results in the formation of quasistationary intermediate states. Nikolai Bazhin Copyright © 2012 Nikolai Bazhin. All rights reserved.