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Several Sequences (several + sequence)
Selected AbstractsActive bacterial community structure along vertical redox gradients in Baltic Sea sedimentENVIRONMENTAL MICROBIOLOGY, Issue 8 2008Anna Edlund Summary Community structures of active bacterial populations were investigated along a vertical redox profile in coastal Baltic Sea sediments by terminal-restriction fragment length polymorphism (T-RFLP) and clone library analysis. According to correspondence analysis of T-RFLP results and sequencing of cloned 16S rRNA genes, the microbial community structures at three redox depths (179, ,64 and ,337 mV) differed significantly. The bacterial communities in the community DNA differed from those in bromodeoxyuridine (BrdU)-labelled DNA, indicating that the growing members of the community that incorporated BrdU were not necessarily the most dominant members. The structures of the actively growing bacterial communities were most strongly correlated to organic carbon followed by total nitrogen and redox potentials. Bacterial identification by sequencing of 16S rRNA genes from clones of BrdU-labelled DNA and DNA from reverse transcription polymerase chain reaction showed that bacterial taxa involved in nitrogen and sulfur cycling were metabolically active along the redox profiles. Several sequences had low similarities to previously detected sequences, indicating that novel lineages of bacteria are present in Baltic Sea sediments. Also, a high number of different 16S rRNA gene sequences representing different phyla were detected at all sampling depths. [source] Disulfide Bond Substitution by Directed Evolution in an Engineered Binding ProteinCHEMBIOCHEM, Issue 8 2009Antoine Drevelle Dr. Abstract Breaking ties: The antitumour protein, neocarzinostatin (NCS), is one of the few drug-carrying proteins used in human therapeutics. However, the presence of disulfide bonds limits this protein's potential development for many applications. This study describes a generic directed-evolution approach starting from NCS-3.24 (shown in the figure complexed with two testosterone molecules) to engineer stable disulfide-free NCS variants suitable for a variety of purposes, including intracellular applications. The chromoprotein neocarzinostatin (NCS) has been intensively studied for its antitumour properties. It has recently been redesigned as a potential drug-carrying scaffold. A potential limit of this protein scaffold, especially for intracellular applications, is the presence of disulfide bonds. The objective of this work was to create a disulfide-free NCS-derived scaffold. A generic targeted approach was developed by using directed evolution methods. As a starting point we used a previously engineered NCS variant in which a hapten binding site had been created. A library was then generated in which cysteine Cys88 and Cys93 and neighbouring residues were randomly substituted. Variants that preserved the hapten binding function were selected by phage display and further screened by colony filtration methods. Several sequences with common features emerged from this process. The corresponding proteins were expressed, purified and their biophysical properties characterised. How these selected sequences rescued folding ability and stability of the disulfide-free protein was carefully examined by using calorimetry and the results were interpreted with molecular simulation techniques. [source] Assessing strategies for improved superfamily recognitionPROTEIN SCIENCE, Issue 7 2005Ian Sillitoe Abstract There are more than 200 completed genomes and over 1 million nonredundant sequences in public repositories. Although the structural data are more sparse (,13,000 nonredundant structures solved to date), several powerful sequence-based methodologies now allow these structures to be mapped onto related regions in a significant proportion of genome sequences. We review a number of publicly available strategies for providing structural annotations for genome sequences, and we describe the protocol adopted to provide CATH structural annotations for completed genomes. In particular, we assess the performance of several sequence-based protocols employing Hidden Markov model (HMM) technologies for superfamily recognition, including a new approach (SAMOSA [sequence augmented models of structure alignments]) that exploits multiple structural alignments from the CATH domain structure database when building the models. Using a data set of remote homologs detected by structure comparison and manually validated in CATH, a single-seed HMM library was able to recognize 76% of the data set. Including the SAMOSA models in the HMM library showed little gain in homolog recognition, although a slight improvement in alignment quality was observed for very remote homologs. However, using an expanded 1D-HMM library, CATH-ISL increased the coverage to 86%. The single-seed HMM library has been used to annotate the protein sequences of 120 genomes from all three major kingdoms, allowing up to 70% of the genes or partial genes to be assigned to CATH superfamilies. It has also been used to recruit sequences from Swiss-Prot and TrEMBL into CATH domain superfamilies, expanding the CATH database eightfold. [source] Origin and geochemistry of Miocene marine evaporites associated with red beds: Great Kavir Basin, Central IranGEOLOGICAL JOURNAL, Issue 1 2007Hossain Rahimpour-Bonab Abstract During the Cenozoic numerous shallow epicontinental evaporite basins formed due to tectonic movements in the Northern Province of the Central Iran Tectonic Zone (the Great Kavir Basin). During the Miocene, due to sea-level fluctuations, thick sequences of evaporites and carbonates accumulated in these basins that subsequently were overlain by continental red beds. Development of halite evaporites with substantial thickness in this area implies inflow of seawater along the narrow continental rift axis. The early ocean basin development was initiated in Early Eocene time and continued up to the Middle Miocene in the isolated failed rift arms. Competition between marine and non-marine environments, at the edge of the encroaching sea, produced several sequences of both abrupt and gradual transition from continental wadi sediments to marginal marine evaporites in the studied area. These evaporites show well-preserved textures indicative of relatively shallow-brine pools. The high Br content of these evaporites indicates marine-derived parent brines that were under the sporadic influence of freshening by meteoric water or replenishing seawater. However, the association of hopper and cornet textures denotes stratified brine that filled a relatively large pool and prevented rapid variations in the Br profile. Unstable basin conditions that triggered modification of parent brine chemistry prevailed in this basin and caused variable distribution patterns for different elements in the chloride units. The presence of sylvite and the absence of Mg-sulphate/chlorides in the paragenetic sequence indicate SO4,depleted parent brine in the studied sequence. Petrographic examinations along with geochemical analyses on these potash-bearing halites reveal parental brines which were a mixture of seawater and CaCl2 -rich brines. The source of CaCl2 -rich brines is ascribed to the presence of local rift systems in the Great Kavir Basin up to the end of the Early Miocene. Copyright © 2007 John Wiley & Sons, Ltd. [source] CHARACTERIZATION OF A DINOFLAGELLATE CRYPTOCHROME BLUE-LIGHT RECEPTOR WITH A POSSIBLE ROLE IN CIRCADIAN CONTROL OF THE CELL CYCLE,JOURNAL OF PHYCOLOGY, Issue 3 2007Stephanie A. Brunelle Karenia brevis (C. C. Davis) G. Hansen et Moestrup is a dinoflagellate responsible for red tides in the Gulf of Mexico. The signaling pathways regulating its cell cycle are of interest because they are the key to the formation of toxic blooms that cause mass marine animal die-offs and human illness. Karenia brevis displays phased cell division, in which cells enter S phase at precise times relative to the onset of light. Here, we demonstrate that a circadian rhythm underlies this behavior and that light quality affects the rate of cell-cycle progression: in blue light, K. brevis entered the S phase early relative to its behavior in white light of similar intensity, whereas in red light, K. brevis was not affected. A data base of 25,000 K. brevis expressed sequence tags (ESTs) revealed several sequences with similarity to cryptochrome blue-light receptors, but none related to known red-light receptors. We characterized the K. brevis cryptochrome (Kb CRY) and modeled its three-dimensional protein structure. Phylogenetic analysis of the photolyase/CRY gene family showed that Kb CRY is a member of the cryptochrome DASH (CRY DASH) clade. Western blotting with an antibody designed to bind a conserved peptide within Kb CRY identified a single band at ,55 kDa. Immunolocalization showed that Kb CRY, like CRY DASH in Arabidopsis, is localized to the chloroplast. This is the first blue-light receptor to be characterized in a dinoflagellate. As the Kb CRY appears to be the only blue-light receptor expressed, it is a likely candidate for circadian entrainment of the cell cycle. [source] Optimal Synthesis of Protein Purification ProcessesBIOTECHNOLOGY PROGRESS, Issue 4 2001Elsa Vásquez-Alvarez There has been an increasing interest in the development of systematic methods for the synthesis of purification steps for biotechnological products, which are often the most difficult and costly stages in a biochemical process. Chromatographic processes are extensively used in the purification of multicomponent biotechnological systems. One of the main challenges in the synthesis of purification processes is the appropriate selection and sequencing of chromatographic steps that are capable of producing the desired product at an acceptable cost and quality. This paper describes mathematical models and solution strategies based on mixed integer linear programming (MILP) for the synthesis of multistep purification processes. First, an optimization model is proposed that uses physicochemical data on a protein mixture, which contains the desired product, to select a sequence of operations with the minimum number of steps from a set of candidate chromatographic techniques that must achieve a specified purity level. Since several sequences that have the minimum number of steps may satisfy the purity level, it is possible to obtain the one that maximizes final purity. Then, a second model that may use the total number of steps obtained in the first model generates a solution with the maximum purity of the product. Whenever the sequence does not affect the final purity or more generally does not impact the objective function, alternative models that are of smaller size are developed for the optimal selection of steps. The models are tested in several examples, containing up to 13 contaminants and a set of 22 candidate high-resolution steps, generating sequences of six operations, and are compared to the current synthesis approaches. [source] | |||||||||||||