Structural and Sequence Characteristics of Long alpha Helices in Globular Proteins

Elucidation of the detailed structural features and sequence requirements for iv helices of various lengths could be very important in understanding secondary structure formation in proteins and, hence. in the protein folding mechanism. An algorithm to characterize the geometry of an alpha helix from its C-alpha coordinates has been developed and used to analyze the structures of long cu helices (number of residues greater than or equal to 25) found in globular proteins, the crystal structure coordinates of which are available from the Brookhaven Protein Data Bank, Ail long a helices can be unambiguously characterized as belonging to one of three classes: linear, curved, or kinked, with a majority being curved. Analysis of the sequences of these helices reveals that the long alpha helices have unique sequence characteristics that distinguish them from the short alpha helices in globular proteins, The distribution and statistical propensities of individual amino acids to occur in long alpha heices are different from those found in short alpha helices, with amino acids having longer side chains and/or having a greater number of functional groups occurring more frequently in these helices, The sequences of the long alpha helices can be correlated with their gross structural features, i.e., whether they are curved, linear, or kinked, and in case of the curved helices, with their curvature.

Serum proteomics of glioma: methods and applications

The prognosis of patients with glioblastoma, the most malignant adult glial brain tumor, remains poor in spite of advances in treatment procedures, including surgical resection, irradiation and chemotherapy.Genetic heterogeneity of glioblastoma warrants extensive studies in order to gain a thorough understanding of the biology of this tumor. While there have been several studies of global transcript profiling of glioma with the identification of gene signatures for diagnosis and disease management, translation into clinics is yet to happen. Serum biomarkers have the potential to revolutionize the process of cancer diagnosis, grading, prognostication and treatment response monitoring. Besides having the advantage that serum can be obtained through a less invasive procedure, it contains molecules at an extraordinary dynamic range of ten orders of magnitude in terms of their concentrations. While the conventional methods, such as 2DE, have been in use for many years, the ability to identify the proteins through mass spectrometry techniques such as MALDI-TOF led to an explosion of interest in proteomics. Relatively new high-throughput proteomics methods such as SELDI-TOF and protein microarrays are expected to hasten the process of serum biomarker discovery. This review will highlight the recent advances in the proteomics platform in discovering serum biomarkers and the current status of glioma serum markers. We aim to provide the principles and potential of the latest proteomic approaches and their applications in the biomarker discovery process. Besides providing a comprehensive list of available serum biomarkers of glioma, we will also propose how these markers will revolutionize the clinical management of glioma patients.

A distinct physiological role of MutY in mutation prevention in mycobacteria

Oxidative damage to DNA results in the occurrence of 7,8-dihydro-B-oxoguanine (8-oxoG) in the genome. In eubacteria, repair of such damage is initiated by two major base-excision repair enzymes, MutM and MutY. We generated a MutY-deficient strain of Mycobacterium smegmatis to investigate the role of this enzyme in DNA repair. The MutY deficiency in M. smegmatis did not result in either a noteworthy susceptibility to oxidative stress or an increase in the mutation rate. However, rifampicin resistant isolates of the MutY-deficient strain showed distinct mutations in the rifampicin-resistance-determining region of rpoB. Besides the expected C to A (or G to T) mutations, an increase in A to C (or T to G) mutations was also observed. Biochemical characterization of mycobacterial MutY (M. smegmatis and M. tuberculosis) revealed an expected excision of A opposite 8-oxoG in DNA. Additionally, excision of G and T opposite 8-oxoG was detected. MutY formed complexes with DNA containing 8-oxoG: A, 8-oxoG: G or 8-oxoG: T but not 8-oxoG : C pairs. Primer extension reactions in cell-free extracts of M. smegmatis suggested error-prone incorporation of nucleotides into the DNA. Based on these observations, we discuss the physiological role of MutY in specific mutation prevention in mycobacteria.

Novel insertion and deletion mutants of RpoB that render Mycobacterium smegmatis RNA polymerase resistant to rifampicin-mediated inhibition of transcription

The startling increase in the occurrence of rifampicin (Rif) resistance in the clinical isolates of Mycobacterium tuberculosis worldwide is posing a serious concern to tuberculosis management. The majority of Rif resistance in bacteria arises from mutations in the RpoB subunit of the RNA polymerase. We isolated M. smegmatis strains harbouring either an insertion (6 aa) or a deletion (10 aa) in their RpoB proteins. Although these strains showed a compromised fitness for growth in 7H9 Middlebrook medium, their resistance to Rif was remarkably high. The attenuated growth of the strains correlated with decreased specific activities of the RNA polymerases from the mutants. While the RNA polymerases from the parent or a mutant strain (harbouring a frequently occurring mutation, H442Y, in RpoB) were susceptible to Rif-mediated inhibition of transcription from calf thymus DNA, those from the insertion and deletion mutants were essentially refractory to such inhibition. Three-dimensional structure modelling revealed that the RpoB amino acids that interact with Rif are either deleted or unable to interact with Rif due to their unsuitable spatial positioning in these mutants. We discuss possible uses of the RpoB mutants in studying transcriptional regulation in mycobacteria and as potential targets for drug design.

Analytical modeling of quantum threshold voltage for triple gate MOSFET

In this work a physically based analytical quantum threshold voltage model for the triple gate long channel metal oxide semiconductor field effect transistor is developed The proposed model is based on the analytical solution of two-dimensional Poisson and two-dimensional Schrodinger equation Proposed model is extended for short channel devices by including semi-empirical correction The impact of effective mass variation with film thicknesses is also discussed using the proposed model All models are fully validated against the professional numerical device simulator for a wide range of device geometries (C) 2010 Elsevier Ltd All rights reserved

Generating synchronizable test sequences based on finite-statemachine with distributed ports

In the area of testing communication systems, the interfaces between systems to be tested and their testers have great impact on test generation and fault detectability. Several types of such interfaces have been standardized by the International Standardization Organization (ISO). A general distributed test architecture, containing distributed interfaces, has been presented in the literature for testing distributed systems based on the Open Distributing Processing (ODP) Basic Reference Model (BRM), which is a generalized version of ISO distributed test architecture. We study in this paper the issue of test selection with respect to such an test architecture. In particular, we consider communication systems that can be modeled by finite state machines with several distributed interfaces, called ports. A test generation method is developed for generating test sequences for such finite state machines, which is based on the idea of synchronizable test sequences. Starting from the initial effort by Sarikaya, a certain amount of work has been done for generating test sequences for finite state machines with respect to the ISO distributed test architecture, all based on the idea of modifying existing test generation methods to generate synchronizable test sequences. However, none studies the fault coverage provided by their methods. We investigate the issue of fault coverage and point out a fact that the methods given in the literature for the distributed test architecture cannot ensure the same fault coverage as the corresponding original testing methods. We also study the limitation of fault detectability in the distributed test architecture.

Stretch effects extracted from inwardly and outwardly propagating spherical premixed flames

Unsteady propagation of spherical flames, both inward and outward, are studied numerically extensively for single-step reaction and for different Lewis numbers of fuel/oxidizer. The dependence of flame speed ratio (s) and flame temperature ratio are obtained for a range of Lewis numbers and stretch (kappa) values. These results of s versus kappa show that the asymptotic theory by Frankel and Sivashinsky is reasonable for outward propagation. Other theories are unsatisfactory both quantitatively and qualitatively. The stretch effects are much higher for negative stretch than for positive stretch, as also seen in the theory of Frankel and Sivashinsky. The linearity of the flame speed ratio vs stretch relationship is restricted to nondimensional stretch of +/-0.1. It is shown further that the results from cylindrical flames are identical to the spherical flame on flame speed ratio versus nondimensional stretch plot thus confirming the generality of the concept of stretch. The comparison of the variation of (ds/dkappa)kappa=0 with beta(Lc - 1) show an offset between the computed and the asymptotic results of Matalon and Matkowsky. The departure of negative stretch results from this variation is significant. Several earlier experimental results are analysed and set out in the form of s versus kappa plot. Comparison of the results with experiments seem reasonable for negative stretch. The results for positive stretch are satisfactory qualitatively for a few cases. For rich propane-air, there are qualitative differences pointing to the need for full chemistry calculations in the extraction of stretch effects.

End-bearing capacity of driven piles in sand using the stress characteristics method: analysis and implementation

The method of stress characteristics has been employed to compute the end-bearing capacity of driven piles. The dependency of the soil internal friction angle on the stress level has been incorporated to achieve more realistic predictions for the end-bearing capacity of piles. The validity of the assumption of the superposition principle while using the bearing capacity equation based on soil plasticity concepts, when applied to deep foundations, has been examined. Fourteen pile case histories were compiled with cone penetration tests (CPT) performed in the vicinity of different pile locations. The end-bearing capacity of the piles was computed using different methods, namely, static analysis, effective stress approach, direct CPT, and the proposed approach. The comparison between predictions made by different methods and measured records shows that the stress-level-based method of stress characteristics compares better with experimental data. Finally, the end-bearing capacity of driven piles in sand was expressed in terms of a general expression with the addition of a new factor that accounts for different factors contributing to the bearing capacity. The influence of the soil nonassociative flow rule has also been included to achieve more realistic results.

Familial Diarrhea Syndrome Caused by an Activating GUCY2C Mutation

BACKGROUND Familial diarrhea disorders are, in most cases, severe and caused by recessive mutations. We describe the cause of a novel dominant disease in 32 members of a Norwegian family. The affected members have chronic diarrhea that is of early onset, is relatively mild, and is associated with increased susceptibility to inflammatory bowel disease, small-bowel obstruction, and esophagitis. METHODS We used linkage analysis, based on arrays with single-nucleotide polymorphisms, to identify a candidate region on chromosome 12 and then sequenced GUCY2C, encoding guanylate cyclase C (GC-C), an intestinal receptor for bacterial heat-stable enterotoxins. We performed exome sequencing of the entire candidate region from three affected family members, to exclude the possibility that mutations in genes other than GUCY2C could cause or contribute to susceptibility to the disease. We carried out functional studies of mutant GC-C using HEK293T cells. RESULTS We identified a heterozygous missense mutation (c.2519G -> T) in GUCY2C in all affected family members and observed no other rare variants in the exons of genes in the candidate region. Exposure of the mutant receptor to its ligands resulted in markedly increased production of cyclic guanosine monophosphate (cGMP). This may cause hyperactivation of the cystic fibrosis transmembrane regulator (CFTR), leading to increased chloride and water secretion from the enterocytes, and may thus explain the chronic diarrhea in the affected family members. CONCLUSIONS Increased GC-C signaling disturbs normal bowel function and appears to have a proinflammatory effect, either through increased chloride secretion or additional effects of elevated cellular cGMP. Further investigation of the relevance of genetic variants affecting the GC-C-CFTR pathway to conditions such as Crohn's disease is warranted. (Funded by Helse Vest Western Norway Regional Health Authority] and the Department of Science and Technology, Government of India.)

Biochemical Properties of MutT2 Proteins from Mycobacterium tuberculosis and M. smegmatis and Their Contrasting Antimutator Roles in Escherichia coli

Mycobacterium tuberculosis, the causative agent of tuberculosis, is at increased risk of accumulating damaged guanine nucleotides such as 8-oxo-dGTP and 8-oxo-GTP because of its residency in the oxidative environment of the host macrophages. By hydrolyzing the oxidized guanine nucleotides before their incorporation into nucleic acids, MutT proteins play a critical role in allowing organisms to avoid their deleterious effects. Mycobacteria possess several MutT proteins. Here, we purified recombinant M. tuberculosis MutT2 (MtuMutT2) and M. smegmatis MutT2 (MsmMutT2) proteins from M. tuberculosis (a slow grower) and M. smegmatis (fast growing model mycobacteria), respectively, for their biochemical characterization. Distinct from the Escherichia coli MutT, which hydrolyzes 8-oxo-dGTP and 8-oxo-GTP, the mycobacterial proteins hydrolyze not only 8-oxo-dGTP and 8-oxo-GTP but also dCTP and 5-methyl-dCTP. Determination of kinetic parameters (K-m and V-max) revealed that while MtuMutT2 hydrolyzes dCTP nearly four times better than it does 8-oxo-dGTP, MsmMutT2 hydrolyzes them nearly equally. Also, MsmMutT2 is about 14 times more efficient than MtuMutT2 in its catalytic activity of hydrolyzing 8-oxo-dGTP. Consistent with these observations, MsmMutT2 but not MtuMutT2 rescues E. coli for MutT deficiency by decreasing both the mutation frequency and A-to-C mutations (a hallmark of MutT deficiency). We discuss these findings in the context of the physiological significance of MutT proteins.

Mycobacterium tuberculosis mutT1 (Rv2985) and ADPRase (Rv1700) proteins constitute a two-stage mechanism of 8-Oxo-dGTP and 8-Oxo-GTP detoxification and adenosine to cytidine mutation avoidance

Approximately one third of the world population is infected with Mycobacterium tuberculosis, the causative agent of tuberculosis. A better understanding of the pathogen biology is crucial to develop new tools/strategies to tackle its spread and treatment. In the host macrophages, the pathogen is exposed to reactive oxygen species, known to damage dGTP and GTP to 8-oxo-dGTP and 8-oxo-GTP, respectively. Incorporation of the damaged nucleotides in nucleic acids is detrimental to organisms. MutT proteins, belonging to a class of Nudix hydrolases, hydrolyze 8-oxo-G nucleoside triphosphates/diphosphates to the corresponding nucleoside monophosphates and sanitize the nucleotide pool. Mycobacteria possess several MutT proteins. However, a functional homolog of Escherichia coli MutT has not been identified. Here, we characterized MtuMutT1 and Rv1700 proteins of M. tuberculosis. Unlike other MutT proteins, MtuMutT1 converts 8-oxo-dGTP to 8-oxo-dGDP, and 8-oxo-GTP to 8-oxo-GDP. Rv1700 then converts them to the corresponding nucleoside monophosphates. This observation suggests the presence of a two-stage mechanism of 8-oxo-dGTP/8-oxo-GTP detoxification in mycobacteria. MtuMutT1 converts 8-oxo-dGTP to 8-oxo-dGDP with a K-m of similar to 50 mu M and V-max of similar to 0.9 pmol/min per ng of protein, and Rv1700 converts 8-oxo-dGDP to 8-oxo-dGMP with a K-m of similar to 9.5 mu M and V-max of similar to 0.04 pmol/min per ng of protein. Together, MtuMutT1 and Rv1700 offer maximal rescue to E. coli for its MutT deficiency by decreasing A to C mutations (a hallmark of MutT deficiency). We suggest that the concerted action of MtuMutT1 and Rv1700 plays a crucial role in survival of bacteria against oxidative stress.

An assessment of terrestrial water storage, rainfall and river discharge over Northern India from satellite data

Terrestrial water storage (TWS) plays a key role in the global water cycle and is highly influenced by climate variability and human activities. In this study, monthly TWS, rainfall and Ganga-Brahmaputra river discharge (GBRD) are analysed over India for the period of 2003-12 using remote sensing satellite data. The spatial pattern of mean TWS shows a decrease over a large and populous region of Northern India comprising the foothills of the Himalayas, the Indo-Gangetic Plains and North East India. Over this region, the mean monthly TWS exhibits a pronounced seasonal cycle and a large interannual variability, highly correlated with rainfall and GBRD variations (r > 0.8) with a lag time of 2 months and 1 month respectively. The time series of monthly TWS shows a consistent and statistically significant decrease of about 1 cm year(-1) over Northern India, which is not associated with changes in rainfall and GBRD. This recent change in TWS suggests a possible impact of rapid industrialization, urbanization and increase in population on land water resources. Our analysis highlights the potential of the Earth-observation satellite data for hydrological applications.

Post-failure Analysis and Fractography of In-plane Tension-Tested Tufted Carbon Fabric-Reinforced Epoxy Composite Laminates

Tufted and plain unidirectional carbon fabric-reinforced epoxy composite laminates were fabricated by vacuum-enhanced resin infusion technology and subjected to in-plane tensile tests with a view to study the changes in mechanical properties and failure responses. Owing to the presence of tufts in the laminates, both the tensile strength and modulus decrease by similar to 38 and similar to 20%, respectively, vis-A -vis the values recorded for plain composites. The fracture features point to the fact that though both the composites fail in brittle manner, they, however, exhibit differing fiber pull out lengths. Further, it was noticed that for the tufted ones, crack originates in the vicinity of tuft thread, spreads through the composite in a brittle manner, and results in a display of shorter fiber pull out lengths. These observations and other results are discussed in this paper.

A unique uracil-DNA binding protein of the uracil DNA glycosylase superfamily

Uracil DNA glycosylases (UDGs) are an important group of DNA repair enzymes, which pioneer the base excision repair pathway by recognizing and excising uracil from DNA. Based on two short conserved sequences (motifs A and B), UDGs have been classified into six families. Here we report a novel UDG, UdgX, from Mycobacterium smegmatis and other organisms. UdgX specifically recognizes uracil in DNA, forms a tight complex stable to sodium dodecyl sulphate, 2-mercaptoethanol, urea and heat treatment, and shows no detectable uracil excision. UdgX shares highest homology to family 4 UDGs possessing Fe-S cluster. UdgX possesses a conserved sequence, KRRIH, which forms a flexible loop playing an important role in its activity. Mutations of H in the KRRIH sequence to S, G, A or Q lead to gain of uracil excision activity in MsmUdgX, establishing it as a novel member of the UDG superfamily. Our observations suggest that UdgX marks the uracil-DNA for its repair by a RecA dependent process. Finally, we observed that the tight binding activity of UdgX is useful in detecting uracils in the genomes.