The Role of UPF0157 in the Folding of M-tuberculosis Dephosphocoenzyme A Kinase and the Regulation of the Latter by CTP

Background: Targeting the biosynthetic pathway of Coenzyme A (CoA) for drug development will compromise multiple cellular functions of the tubercular pathogen simultaneously. Structural divergence in the organization of the penultimate and final enzymes of CoA biosynthesis in the host and pathogen and the differences in their regulation mark out the final enzyme, dephosphocoenzyme A kinase (CoaE) as a potential drug target. Methodology/Principal Findings: We report here a complete biochemical and biophysical characterization of the M. tuberculosis CoaE, an enzyme essential for the pathogen's survival, elucidating for the first time the interactions of a dephosphocoenzyme A kinase with its substrates, dephosphocoenzyme A and ATP; its product, CoA and an intrinsic yet novel inhibitor, CTP, which helps modulate the enzyme's kinetic capabilities providing interesting insights into the regulation of CoaE activity. We show that the mycobacterial enzyme is almost 21 times more catalytically proficient than its counterparts in other prokaryotes. ITC measurements illustrate that the enzyme follows an ordered mechanism of substrate addition with DCoA as the leading substrate and ATP following in tow. Kinetic and ITC experiments demonstrate that though CTP binds strongly to the enzyme, it is unable to participate in DCoA phosphorylation. We report that CTP actually inhibits the enzyme by decreasing its Vmax. Not surprisingly, a structural homology search for the modeled mycobacterial CoaE picks up cytidylmonophosphate kinases, deoxycytidine kinases, and cytidylate kinases as close homologs. Docking of DCoA and CTP to CoaE shows that both ligands bind at the same site, their interactions being stabilized by 26 and 28 hydrogen bonds respectively. We have also assigned a role for the universal Unknown Protein Family 0157 (UPF0157) domain in the mycobacterial CoaE in the proper folding of the full length enzyme. Conclusions/Significance: In view of the evidence presented, it is imperative to assign a greater role to the last enzyme of Coenzyme A biosynthesis in metabolite flow regulation through this critical biosynthetic pathway.

Functional diversity of human protein kinase splice variants marks significant expansion of human kinome

Background: Protein kinases are involved in diverse spectrum of cellular processes. Availability of draft version of the human genomic data in the year 2001 enabled recognition of repertoire of protein kinases. However, over the years the human genomic data is being refined and the current release of human genomic data has helped us to recognize a larger repertoire of over 900 human protein kinases represented mainly by splice variants. Results: Many of these identified protein kinases are alternatively spliced products. Interestingly, some of the human kinase splice variants appear to be significantly diverged in terms of their functional properties as represented by incorporation or absence of one or more domains. Many sets of protein kinase splice variants have substantially different domain organization and in a few sets of splice variants kinase domains belong to different subfamilies of kinases suggesting potential participation in different signal transduction pathways. Conclusions: Addition or deletion of a domain between splice variants of multi-domain kinases appears to be a means of generating differences in the functional features of otherwise similar kinases. It is intriguing that marked sequence diversity within the catalytic regions of some of the splice variant kinases result in kinases belonging to different subfamilies. These human kinase splice variants with different functions might contribute to diversity of eukaryotic cellular signaling.

Analysis of the MAP/G(a,b)/1/N queue with multiple vacations

This paper deals with a batch service queue and multiple vacations. The system consists of a single server and a waiting room of finite capacity. Arrival of customers follows a Markovian arrival process (MAP). The server is unavailable for occasional intervals of time called vacations, and when it is available, customers are served in batches of maximum size ‘b’ with a minimum threshold value ‘a’. We obtain the queue length distributions at various epochs along with some key performance measures. Finally, some numerical results have been presented.

Characterization of mouse neuronal Ca2+/calmodulin kinase II inhibitor alpha

We have overexpressed an 8.5-kDa mouse Ca2+/calmodulin kinase II inhibitor a protein (mCaMKIIN alpha) in Escherichia coli and demonstrate that the recombinant protein is a potent inhibitor of Ca2+/calmodulin kinase 11 (CaMKII) in vitro. However, antibodies raised against recombinant mCaMKIIN alpha. react with an similar to 37-kDa protein present in mouse brain. The pattern of expression of the similar to 37-kDa protein is similar to that of mCaMKIIN alpha mRNA as both are expressed in normal but not Japanese encephalitis virus (JEV)-infected mouse brain. Subcellular localization studies indicate that the similar to 37-kDa protein is present in the post-synaptic density (PSD) where mCaMKII alpha is known to perform key regulatory functions. We conclude that the similar to 37-kDa protein identified in this study is mCaMKIIN alpha. and its localization in the PSD indicates a novel role for this protein in the regulation of neuronal CaMKII alpha. (c) 2007 Elsevier B.V. All rights reserved.

Critical lattice size limit for synchronized chaotic state in one- and two-dimensional diffusively coupled map lattices

We consider diffusively coupled map lattices with P neighbors (where P is arbitrary) and study the stability of the synchronized state. We show that there exists a critical lattice size beyond which the synchronized state is unstable. This generalizes earlier results for nearest neighbor coupling. We confirm the analytical results by performing numerical simulations on coupled map lattices with logistic map at each node. The above analysis is also extended to two-dimensional P-neighbor diffusively coupled map lattices.

Hot deformation behaviour of Mg–3Al alloy—A study using processing map

The hot deformation behaviour of Mg–3Al alloy has been studied using the processing-map technique. Compression tests were conducted in the temperature range 250–550 °C and strain rate range 3 × 10−4 to 102 s−1 and the flow stress data obtained from the tests were used to develop the processing map. The various domains in the map corresponding to different dissipative characteristics have been identified as follows: (i) grain boundary sliding (GBS) domain accommodated by slip controlled by grain boundary diffusion at slow strain-rates (<10−3 s−1) in the temperature range from 350 to 450 °C, (ii) two different dynamic recrystallization (DRX) domains with a peak efficiency of 42% at 550 °C/10−1 s−1 and 425 °C/102 s−1 governed by stress-assisted cross-slip and thermally activated climb as the respective rate controlling mechanisms and (iii) dynamic recovery (DRV) domain below 300 °C in the intermediate strain rate range from 3 × 10−2 to 3 × 10−1 s−1. The regimes of flow instability have also been delineated in the processing map using an instability criterion. Adiabatic shear banding at higher strain rates (>101 s−1) and solute drag by substitutional Al atoms at intermediate strain rates (3 × 10−2 to 3 × 10−1 s−1) in the temperature range (350–450 °C) are responsible for flow instability. The relevance of these mechanisms with reference to hot working practice of the material has been indicated. The processing maps of Mg–3Al alloy and as-cast Mg have been compared qualitatively to elucidate the effect of alloying with aluminum on the deformation behaviour of magnesium.

Quantitative photoacoustic tomography from boundary pressure measurements: noniterative recovery of optical absorption coefficient from the reconstructed absorbed energy map

We describe a noniterative method for recovering optical absorption coefficient distribution from the absorbed energy map reconstructed using simulated and noisy boundary pressure measurements. The source reconstruction problem is first solved for the absorbed energy map corresponding to single- and multiple-source illuminations from the side of the imaging plane. It is shown that the absorbed energy map and the absorption coefficient distribution, recovered from the single-source illumination with a large variation in photon flux distribution, have signal-to-noise ratios comparable to those of the reconstructed parameters from a more uniform photon density distribution corresponding to multiple-source illuminations. The absorbed energy map is input as absorption coefficient times photon flux in the time-independent diffusion equation (DE) governing photon transport to recover the photon flux in a single step. The recovered photon flux is used to compute the optical absorption coefficient distribution from the absorbed energy map. In the absence of experimental data, we obtain the boundary measurements through Monte Carlo simulations, and we attempt to address the possible limitations of the DE model in the overall reconstruction procedure.

Specific Small-Molecule Activator of Aurora Kinase A Induces Autophosphorylation in a Cell-Free System

Aurora kinases are essential for chromosomal segregation and cell division and thereby important for maintaining the proper genomic integrity. There are three classes of aurora kinases in humans: A, B, and C. Aurora kinase A is frequently overexpressed in various cancers. The link of the overexpression and tumorigenesis is yet to be understood. By employing virtual screening, we have found that anacardic acid, a pentadecane aliphatic chain containing hydroxylcarboxylic acid, from cashew nut shell liquid could be docked in Aurora kinases A and B. Remarkably, we found that anacardic acid could potently activate the Aurora kinase A mediated phosphorylation of histone H3, but at a similar concentration the activity of aurora kinase B remained unaffected in vitro. Mechanistically, anacardic acid induces the structural changes and also the autophosphorylation of the aurora kinase A to enhance the enzyme activity. This data thus indicate anacardic acid as the first small-molecule activator of Aurora kinase, which could be highly useful for probing the function of hyperactive (overexpressed) Aurora kinase A.

Mutational analysis of the insulin‑like growth factor 1 receptor tyrosine kinase domain in non-small cell lung cancer patients

The insulin‑like growth factor 1 receptor (IGF1R) pathway plays an important role in the pathogenesis of non‑small cell lung cancer (NSCLC) and also provides a mechanism of resistance to targeted therapies. IGF1R is therefore an ideal therapeutic target and several inhibitors have entered clinical trials. However, thus far the response to these inhibitors has been poor, highlighting the importance of predictive biomarkers to identify patient cohorts who will benefit from these targeted agents. It is well‑documented that mutations and/or deletions in the epidermal growth factor receptor (EGFR) tyrosine kinase (TK) domain predict sensitivity of NSCLC patients to EGFR TK inhibitors. Single‑nucleotide polymorphisms (SNPs) in the IGF pathway have been associated with disease, including breast and prostate cancer. The aim of the present study was to elucidate whether the IGF1R TK domain harbours SNPs, somatic mutations or deletions in NSCLC patients and correlates the mutation status to patient clinicopathological data and prognosis. Initially 100 NSCLC patients were screened for mutations/deletions in the IGF1R TK domain (exons 16‑21) by sequencing analysis. Following the identification of SNP rs2229765, a further 98 NSCLC patients and 866 healthy disease‑free control patients were genotyped using an SNP assay. The synonymous SNP (rs2229765) was the only aberrant base change identified in the IGF1R TK domain of 100 NSCLC patients initially analysed. SNP rs2229765 was detected in exon 16 and was found to have no significant association between IGF1R expression and survival. The GA genotype was identified in 53.5 and 49.4% of NSCLC patients and control individuals, respectively. No significant difference was found in the genotype (P=0.5487) or allele (P=0.9082) frequencies between the case and control group. The present findings indicate that in contrast to the EGFR TK domain, the IGF1R TK domain is not frequently mutated in NSCLC patients. The synonymous SNP (rs2229765) had no significant association between IGF1R expression and survival in the cohort of NSCLC patients.

Revisiting Coptotermes (Isoptera: Rhinotermitidae): a global taxonomic road map for species validity and distribution of an economically important subterranean termite genus

Coptotermes Wasmann (Isoptera: Rhinotermitidae) is one of the most economically important subterranean termite genera and some species are successful invaders. However, despite its important pest status, the taxonomic validity of many named Coptotermes species remains unclear. In this study, we reviewed all named species within the genus and investigated evidence supporting the validity of each named species. Species were systematically scrutinized according to the region of their original description: Southeast Asia, India, China, Africa, the Neotropics, and Australia. We estimate that of the currently 69 named species described by accepted nomenclatural rules, only 21 taxa have solid evidence for validity, 44 names have uncertain status, and the remaining species names should be synonymized or were made unavailable. Species with high degrees of invasiveness may be known under additional junior synonyms due to independent parochial descriptions. Molecular data for a vast majority of species are scarce and significant effort is needed to complete the taxonomic and phylogenetic revision of the genus. Because of the wide distribution of Coptotermes, we advocate for an integrative taxonomic effort to establish the distribution of each putative species, provide specimens and corresponding molecular data, check original descriptions and type specimens (if available), and provide evidence for a more robust phylogenetic position of each species. This study embodies both consensus and contention of those studying Coptotermes and thus pinpoints the current uncertainty of many species. This project is intended to be a roadmap for identifying those Coptotermes species names that need to be more thoroughly investigated, as an incentive to complete a necessary revision process.

Analysis of connectivity map: Control to glutamate injured and phenobarbital treated neuronal network

We study the responses of a cultured neural network when it is exposed to epileptogenesis glutamate injury causing epilepsy and subsequent treatment with phenobarbital by constructing connectivity map of neurons using correlation matrix. This study is particularly useful in understanding the pharmaceutical drug induced changes in the neuronal network properties with insights into changes at the systems biology level. (C) 2010 American Institute of Physics. [doi:10.1063/1.3398025]

Essential role of PI3-kinase pathway in p53-mediated transcription: Implications in cancer chemotherapy

The PI3-kinase pathway is the target of inactivation in achieving better cancer chemotherapy. Here, we report that p53-mediated transcription is inhibited by pharmacological inhibitors and a dominant-negative mutant of PI3-kinase, and this inhibition was relieved by a constitutively active mutant of PI3-kinase. Akt/PKB and mTOR, the downstream effectors of PI3-kinase, were also found to be essential. LY294002 (PI3-kinase inhibitor) pre-treatment altered the post-translational modifications and the sub-cellular localization of p53. Although LY294002 increased the chemosensitivity of cells to low concentrations of adriamycin (adriamycin-low), it protected the cells from cytotoxicity induced by high concentrations of adriamycin (adriamycin-high) in a p53-dependent manner. Further, we found that LY294002 completely abolished the activation of p53 target genes (particularly pro-apoptotic) under adriamycin-high conditions, whereas it only marginally repressed the p53 target genes under adriamycin-low conditions; in fact, it further activated the transcription of NOXA, HRK, APAF1 and CASP5 genes. Thus, the differential effect of PI3-kinase on p53 functions seems to be responsible for the differential regulation of DNA damage-induced cytotoxicity and cell death by PI3-kinase. Our finding becomes relevant in the light of ongoing combination chemotherapy trials with the PI3-kinase pathway inhibitors and underscores the importance of p53 status in the careful formulation of combination chemotherapies. Oncogene (2010) 29, 3605-3618; doi: 10.1038/onc.2010.123; published online 26 April 2010

M.tuberculosis Pantothenate Kinase: Dual Substrate Specificity and Unusual Changes in Ligand Locations

Kinetic measurements of enzyme activity indicate that type I pantothenate kinase from Mycobacterium tuberculosis has dual substrate specificity for ATP and GTP, unlike the enzyme from Escherichia coli, which shows a higher specificity for ATP. A molecular explanation for the difference in the specificities of the two homologous enzymes is provided by the crystal structures of the complexes of the M. tuberculosis enzyme with (1) GMPPCP and pantothenate, (2) GDP and phosphopantothenate, (3) GDP, (4) GDP and pantothenate, (5) AMPPCP, and (6) GMPPCP, reported here, and the structures of the complexes of the two enzymes involving coenzyme A and different adenyl nucleotides reported earlier. The explanation is substantially based on two critical substitutions in the amino acid sequence and the local conformational change resulting from them. The structures also provide a rationale for the movement of ligands during the action of the mycobacterial enzyme. Dual specificity of the type exhibited by this enzyme is rare. The change in locations of ligands during action,observed in the case of the M. tuberculosis enzyme, is unusual, so is the striking difference between two homologous enzymes in the geometryof the binding site, locations of ligands, and specificity. Furthermore, the dual specificity of the mycobacterial enzyme appears to have been caused by a biological necessity. (C) 2010 Elsevier Ltd.All rights reserved.

Low-Complexity Near-MAP Decoding of Large Non-Orthogonal STBCs Using PDA

Non-orthogonal space-time block codes (STBC) from cyclic division algebras (CDA) are attractive because they can simultaneously achieve both high spectral efficiencies (same spectral efficiency as in V-BLAST for a given number of transmit antennas) as well as full transmit diversity. Decoding of non-orthogonal STBCs with hundreds of dimensions has been a challenge. In this paper, we present a probabilistic data association (PDA) based algorithm for decoding non-orthogonal STBCs with large dimensions. Our simulation results show that the proposed PDA-based algorithm achieves near SISO AWGN uncoded BER as well as near-capacity coded BER (within 5 dB of the theoretical capacity) for large non-orthogonal STBCs from CDA. We study the effect of spatial correlation on the BER, and show that the performance loss due to spatial correlation can be alleviated by providing more receive spatial dimensions. We report good BER performance when a training-based iterative decoding/channel estimation is used (instead of assuming perfect channel knowledge) in channels with large coherence times. A comparison of the performances of the PDA algorithm and the likelihood ascent search (LAS) algorithm (reported in our recent work) is also presented.