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At least 19 records

Catalysis and biocatalysis program

The annual report presents the fiscal year (FY) 1990 research activities and accomplishments for the Catalysis and Biocatalysis Program of the Advanced Industrial Concepts Division (AICD), Office of Industrial Technologies of the Department of Energy (DOE). The mission of the AICD is to create a balanced program of high risk, long term, directed interdisciplinary research and development that will improve energy efficiency and enhance fuel flexibility in the industrial sector. The Catalysis and Biocatalysis Program's technical activities were organized into five work elements: the Molecular Modeling and Catalysis by Design element; the Applied Microbiology and Genetics element; the Bioprocess Engineering element; the Separations and Novel Chemical Processes element; and the Process Design and Analysis element.

Source record↗

On the early emergence of reverse transcription: theoretical basis and experimental evidence

Reverse transcriptase (RT) was first discovered as an essential catalyst in the biological cycle of retroviruses. However, in the past years evidence has accumulated showing that RTs are involved in a surprisingly large number of RNA-mediated transpositional events that include both viral and nonviral genetic entities. Although it is probable that some RT-bearing genetic elements like the different types of AIDS viruses and the mammalian LINE family have arisen in recent geological times, the possibility that reverse transcription first took place in the early Archean is supported by (1) the hypothesis that RNA preceded DNA as cellular genetic material; (2) the existence of homologous regions of the subunit tau of the E. coli DNA polymerase III with the simian immunodeficiency virus RT, the hepatitis B virus RT, and the beta' subunit of the E. coli RNA polymerase (McHenry et al. 1988); (3) the presence of several conserved motifs, including a 14-amino-acid segment that consists of an Asp-Asp pair flanked by hydrophobic amino acids, which are found in all RTs and in most cellular and viral RNA polymerases. However, whether extant RTs descend from the primitive polymerase involved in the RNA-to-DNA transition remains unproven. Substrate specificity of the AMV and HIV-1 RTs can be modified in the presence of Mn2+, a cation which allows them to add ribonucleotides to an oligo (dG) primer in a template-dependent reaction. This change in specificity is comparable to that observed under similar conditions in other nucleic acid polymerases. This experimentally induced change in RT substrate specificity may explain previous observations on the misincorporation of ribonucleotides by the Maloney murine sarcoma virus RT in the minus and plus DNA of this retrovirus (Chen and Temin 1980). Our results also suggest that HIV-infected macrophages and T-cell cells may contain mixed polynucleotides containing both ribo- and deoxyribonucleotides. The evolutionary significance of these changes in substrate specificities of nucleic acid polymerases is also discussed.

Non-NASA Center↗

Ames interactive molecular model building system - A 3-D computer modelling system applied to the study of the origin of life

The investigation of specific interactions among biological molecules must take into consideration the stereochemistry of the structures. Thus, models of the molecules are essential for describing the spatial organization of potentially interacting groups, and estimations of conformation are required for a description of spatial organization. Both the function of visualizing molecules, and that of estimating conformation through calculations of energy, are part of the molecular modeling system described in the present paper. The potential uses of the system in investigating some aspects of the origin of life rest on the assumption that translation of conformation from genetic elements to catalytic elements would have been required for the development of the first replicating systems subject to the process of biological evolution.

Coeckelenbergh, Y.↗

Phenotypic characterization of xpr, a global regulator of extracellular virulence factors in Staphylococcus aureus

We recently described a Tn551 insertion in the chromosome of Staphylococcus aureus S6C that resulted in drastically reduced expression of extracellular lipase (M. S. Smeltzer, S. R. Gill, and J. J. Iandolo, J. Bacteriol. 174:4000-4006, 1992). The insertion was localized to a chromosomal site (designated omega 1058) distinct from the lipase structural gene (geh) and the accessory gene regulator (agr), both of which were structurally intact in the lipase-negative (Lip-) mutants. In this report, we describe a phenotypic comparison between strains S6C, a hyperproducer of enterotoxin B; KSI9051, a derivative of S6C carrying the Tn551 insertion at omega 1058; ISP546, an 8325-4 strain that carries a Tn551 insertion in the agr locus; and ISP479C, the parent strain of ISP546 cured of the Tn551 delivery plasmid pI258repA36. Compared with their respective parent strains, ISP546 and KSI9051 produced greatly reduced amounts of lipase, alpha-toxin, delta-toxin, protease, and nuclease. KSI9051 also produced reduced amounts of staphylococcal enterotoxin B. Coagulase production was increased in ISP546 but not in KSI9051. Using a mouse model, we also demonstrated that ISP546 and KSI9051 were far less virulent than ISP479C and S6C. We have designated the genetic element defined by the Tn551 insertion at omega 1058 xpr to denote its role as a regulator of extracellular protein synthesis. We conclude that xpr and agr are similar and possibly interactive regulatory genes that play an important role in pathogenesis of staphylococcal disease.

NASA Discipline Cell Biology↗

Nasa GeneLab Computomics Reveal Horizontal Gene Transfer on International Space Station Environmental Metagenomes

Prokaryotic lifeforms can be observed to demonstrate many keen adaptive advantages, perhaps facilitated by a nature simplistic relative to divergent domains of life. In particular, decompartmentalized gene expression facilitates adaptation by allowing free exchange of genetic material, albeit at the cost of increased susceptibility to genetic damage. Thus, these lifeforms must compensate by embracing diverse investment strategies in an attempt to “brute force” the evolvability equation through precipitous genesis, lean metabolic efficiency, and sheer population. This prokaryotic archetype also enables symbiotic relationships with secondary mobile genetic elements known as plasmids, which have been shown to drive evolution on rapid temporal scales through processes such as conjugation and transformation. This study attempts to decipher whether these mechanisms of horizontal gene transfer (HGT) are major factors in determining prokaryote fitness within a unique isolated environment, the International Space Station (ISS). The ISS Microbial Tracking (MT) project has generated a wealth of data concerning the successive reigns of microbial genera that appear to thrive amidst harsh conditions for life. Despite relatively higher doses of ionizing radiation as compared to Earth, complications associated with microgravity, and the anti-microbial mélange deployed, microbial life still persists in this environment. The NASA GeneLab serves as a data repository and analysis platform to enable researchers to access space flight factor related data. With the use of GeneLab’s modern computational suites (computomics), phylogenetic and functional genomic investigations of HGT events were conducted on the data generated from the MT-1 project. The putative data concerning the plasmid population (plasmidome) of the ISS was algorithmically derived and compared to those of habitats with similar environmental dynamics- such as living quarters and hospitals- to investigate whether these HGT elements may play crucial role(s) in shaping the microbiome of this closed habitat that serves as the only inhabited structure in space.

Bense, Nicholas↗

ECUT (Energy Conversion and Utilization Technologies) program: Biocatalysis Project

Fiscal year 1987 research activities and accomplishments for the Biocatalysis Project of the U.S. Department of Energy, Energy Conversion and Utilization Technologies (ECUT) Division are presented. The project's technical activities were organized into three work elements. The Molecular Modeling and Applied Genetics work element includes modeling and simulation studies to verify a dynamic model of the enzyme carboxypeptidase; plasmid stabilization by chromosomal integration; growth and stability characteristics of plasmid-containing cells; and determination of optional production parameters for hyper-production of polyphenol oxidase. The Bioprocess Engineering work element supports efforts in novel bioreactor concepts that are likely to lead to substantially higher levels of reactor productivity, product yields, and lower separation energetics. The Bioprocess Design and Assessment work element attempts to develop procedures (via user-friendly computer software) for assessing the economics and energetics of a given biocatalyst process.

Source record↗

ECUT (Energy Conversion and Utilization Technologies) program: Biocatalysis project

The Annual Report presents the fiscal year (FY) 1988 research activities and accomplishments, for the Biocatalysis Project of the U.S. Department of Energy, Energy Conversion and Utilization Technologies (ECUT) Division. The ECUT Biocatalysis Project is managed by the Jet Propulsion Laboratory, California Institute of Technology. The Biocatalysis Project is a mission-oriented, applied research and exploratory development activity directed toward resolution of the major generic technical barriers that impede the development of biologically catalyzed commercial chemical production. The approach toward achieving project objectives involves an integrated participation of universities, industrial companies and government research laboratories. The Project's technical activities were organized into three work elements: (1) The Molecular Modeling and Applied Genetics work element includes research on modeling of biological systems, developing rigorous methods for the prediction of three-dimensional (tertiary) protein structure from the amino acid sequence (primary structure) for designing new biocatalysis, defining kinetic models of biocatalyst reactivity, and developing genetically engineered solutions to the generic technical barriers that preclude widespread application of biocatalysis. (2) The Bioprocess Engineering work element supports efforts in novel bioreactor concepts that are likely to lead to substantially higher levels of reactor productivity, product yields and lower separation energetics. Results of work within this work element will be used to establish the technical feasibility of critical bioprocess monitoring and control subsystems. (3) The Bioprocess Design and Assessment work element attempts to develop procedures (via user-friendly computer software) for assessing the energy-economics of biocatalyzed chemical production processes, and initiation of technology transfer for advanced bioprocesses.

Baresi, Larry↗

Distinct transcripts are recognized by sense and antisense riboprobes for a member of the murine HSP70 gene family, HSP70.2, in various reproductive tissues

The expression of hsp70.2, an hsp70 gene family member, originally characterized by its high levels of expression in germ cells in the adult mouse testis, was detected in several other reproductive tissues, including epididymis, prostate, and seminal vesicles, as well as in extraembryonic tissues of mid-gestation fetuses. In addition, hybridization with RNA probes transcribed in the sense orientation surprisingly indicated the presence of slightly larger "antisense" transcripts in several tissues. The levels of antisense transcripts varied among the tissues, with the highest signal detected in the prostate and no signal being detectable in the testis. Consistent with these results, in situ hybridization analysis clearly localized the sense-orientation transcripts to pachytene spermatocytes, while no antisense-orientation transcripts were observed in adjacent sections of the same tubules. Our findings have thus shown that although hsp70.2 was expressed abundantly and in a highly stage-specific manner in the male germ line, it was also expressed in other murine tissues. Furthermore, we have made the surprising observation of antisense transcription of the hsp70.2 gene in several mouse tissues, revealing another level of complexity in the regulation and function of heat shock proteins.

Non-NASA Center↗

ECUT (Energy Conversion and Utilization Technologies Program). Biocatalysis Project

Presented are the FY 1985 accomplishments, activities, and planned research efforts of the Biocatalysis Project of the U.S. Department of Energy, Energy Conversion and Utilization Technologies (ECUT) Program. The Project's technical activities were organized as follows: In the Molecular Modeling and Applied Genetics work element, research focused on (1) modeling and simulation studies to establish the physiological basis of high temperature tolerance in a selected enzyme and the catalytic mechanisms of three species of another enzyme, and (2) determining the degree of plasmid amplification and stability of several DNA bacterial strains. In the Bioprocess Engineering work element, research focused on (1) studies of plasmid propagation and the generation of models, (2) developing methods for preparing immobilized biocatalyst beads, and (3) developing an enzyme encapsulation method. In the Process Design and Analysis work element, research focused on (1) further refinement of a test case simulation of the economics and energy efficiency of alternative biocatalyzed production processes, (2) developing a candidate bioprocess to determine the potential for reduced energy consumption and facility/operating costs, and (3) a techno-economic assessment of potential advancements in microbial ammonia production.

Source record↗

Time- and dose-related interactions between glucocorticoid and cyclic adenosine 3',5'-monophosphate on CCAAT/enhancer-binding protein-dependent insulin-like growth factor I expression by osteoblasts

Glucocorticoid has complex effects on osteoblasts. Several of these changes appear to be related to steroid concentration, duration of exposure, or specific effects on growth factor expression or activity within bone. One important bone growth factor, insulin-like growth factor I (IGF-I), is induced in osteoblasts by hormones such as PGE2 that increase intracellular cAMP levels. In this way, PGE2 activates transcription factor CCAAT/enhancer-binding protein-delta (C/EBPdelta) and enhances its binding to a specific control element found in exon 1 in the IGF-I gene. Our current studies show that preexposure to glucocorticoid enhanced C/EBPdelta and C/EBPbeta expression by osteoblasts and thereby potentiated IGF-I gene promoter activation in response to PGE2. Importantly, this directly contrasts with inhibitory effects on IGF-I expression that result from sustained or pharmacologically high levels of glucocorticoid exposure. Consistent with the stimulatory effect of IGF-I on bone protein synthesis, pretreatment with glucocorticoid sensitized osteoblasts to PGE2, and in this context significantly enhanced new collagen and noncollagen protein synthesis. Therefore, pharmacological levels of glucocorticoid may reduce IGF-I expression by osteoblasts and cause osteopenic disease, whereas physiological transient increases in glucocorticoid may permit or amplify the effectiveness of hormones that regulate skeletal tissue integrity. These events appear to converge on the important role of C/EBPdelta and C/EBPbeta on IGF-I expression by osteoblasts.

NASA Program Biomedical Research and Countermeasur↗

The Toll pathway is required in the epidermis for muscle development in the Drosophila embryo

The Toll signaling pathway functions in several Drosophila processes, including dorsal-ventral pattern formation and the immune response. Here, we demonstrate that this pathway is required in the epidermis for proper muscle development. Previously, we showed that the zygotic Toll protein is necessary for normal muscle development; in the absence of zygotic Toll, close to 50% of hemisegments have muscle patterning defects consisting of missing, duplicated and misinserted muscle fibers (Halfon, M.S., Hashimoto, C., and Keshishian, H., Dev. Biol. 169, 151-167, 1995). We have now also analyzed the requirements for easter, spatzle, tube, and pelle, all of which function in the Toll-mediated dorsal-ventral patterning pathway. We find that spatzle, tube, and pelle, but not easter, are necessary for muscle development. Mutations in these genes give a phenotype identical to that seen in Toll mutants, suggesting that elements of the same pathway used for Toll signaling in dorsal-ventral development are used during muscle development. By expressing the Toll cDNA under the control of distinct Toll enhancer elements in Toll mutant flies, we have examined the spatial requirements for Toll expression during muscle development. Expression of Toll in a subset of epidermal cells that includes the epidermal muscle attachment cells, but not Toll expression in the musculature, is necessary for proper muscle development. Our results suggest that signals received by the epidermis early during muscle development are an important part of the muscle patterning process.

Non-NASA Center↗

B-DNA to Z-DNA structural transitions in the SV40 enhancer: stabilization of Z-DNA in negatively supercoiled DNA minicircles

During replication and transcription, the SV40 control region is subjected to significant levels of DNA unwinding. There are three, alternating purine-pyrimidine tracts within this region that can adopt the Z-DNA conformation in response to negative superhelix density: a single copy of ACACACAT and two copies of ATGCATGC. Since the control region is essential for both efficient transcription and replication, B-DNA to Z-DNA transitions in these vital sequence tracts may have significant biological consequences. We have synthesized DNA minicircles to detect B-DNA to Z-DNA transitions in the SV40 enhancer, and to determine the negative superhelix density required to stabilize the Z-DNA. A variety of DNA sequences, including the entire SV40 enhancer and the two segments of the enhancer with alternating purine-pyrimidine tracts, were incorporated into topologically relaxed minicircles. Negative supercoils were generated, and the resulting topoisomers were resolved by electrophoresis. Using an anti-Z-DNA Fab and an electrophoretic mobility shift assay, Z-DNA was detected in the enhancer-containing minicircles at a superhelix density of -0.05. Fab saturation binding experiments demonstrated that three, independent Z-DNA tracts were stabilized in the supercoiled minicircles. Two other minicircles, each with one of the two alternating purine-pyrimidine tracts, also contained single Z-DNA sites. These results confirm the identities of the Z-DNA-forming sequences within the control region. Moreover, the B-DNA to Z-DNA transitions were detected at superhelix densities observed during normal replication and transcription processes in the SV40 life cycle.

Non-NASA Center↗

Trace elements in Antarctic meteorites: Weathering and genetic information

Antarctic meteorite discoveries have created great scientific interest due to the large number of specimens recovered (approximately 7000) and because included are representatives of hitherto rare or unknown types. Antarctic meteorites are abundant because they have fallen over long periods and were preserved, transported, and concentrated by the ice sheets. The weathering effects on the Antarctic meteorites are described. Weathering effects of trace element contents of H5 chondrites were studied in detail. The results are examined. The properties of Antarctic finds and non-Antarctic falls are discussed.

Lipschutz, M. E.↗

Trace element distribution in mineral separates of the Allende inclusions and their genetic implications

Concentrations of the rare earth elements (REE), Sc, Co, Fe, Zn, Ir, Na, and Cr were determined for mineral separates of the coarseand fine-grained types (group I and II) of the Allende inclusions. These data in combination with other data suggest that the minerals in the coarse-grained inclusions (group I) crystallized in a closed system with respect to refractory elements although a totally molten stage is precluded. The data also indicate that fine-grained (group II) inclusions were formed by condensation from a super-cooled nebular gas; REE-rich clinopyroxene and spinel were formed earlier than REE-poor sodalite and nepheline. In addition, pre-existing Mg isotope anomalies in the coarse-grained inclusions must have been erased during the heating stage.

Nagasawa, H.↗

Saccharomyces cerevisiae gene expression changes during rotating wall vessel suspension culture

This study utilizes Saccharomyces cerevisiae to study genetic responses to suspension culture. The suspension culture system used in this study is the high-aspect-ratio vessel, one type of the rotating wall vessel, that provides a high rate of gas exchange necessary for rapidly dividing cells. Cells were grown in the high-aspect-ratio vessel, and DNA microarray and metabolic analyses were used to determine the resulting changes in yeast gene expression. A significant number of genes were found to be up- or downregulated by at least twofold as a result of rotational growth. By using Gibbs promoter alignment, clusters of genes were examined for promoter elements mediating these genetic changes. Candidate binding motifs similar to the Rap1p binding site and the stress-responsive element were identified in the promoter regions of differentially regulated genes. This study shows that, as in higher order organisms, S. cerevisiae changes gene expression in response to rotational culture and also provides clues for investigations into the signaling pathways involved in gravitational response.

Non-NASA Center↗

Mobile transporter path planning

The use of a genetic algorithm (GA) for solving the mobile transporter path planning problem is investigated. The mobile transporter is a traveling robotic vehicle proposed for the space station which must be able to reach any point of the structure autonomously. Elements of the genetic algorithm are explored in both a theoretical and experimental sense. Specifically, double crossover, greedy crossover, and tournament selection techniques are examined. Additionally, the use of local optimization techniques working in concert with the GA are also explored. Recent developments in genetic algorithm theory are shown to be particularly effective in a path planning problem domain, though problem areas can be cited which require more research.

Baffes, Paul↗

Mobile transporter path planning using a genetic algorithm approach

The use of an optimization technique known as a genetic algorithm for solving the mobile transporter path planning problem is investigated. The mobile transporter is a traveling robotic vehicle proposed for the Space Station which must be able to reach any point of the structure autonomously. Specific elements of the genetic algorithm are explored in both a theoretical and experimental sense. Recent developments in genetic algorithm theory are shown to be particularly effective in a path planning problem domain, though problem areas can be cited which require more research. However, trajectory planning problems are common in space systems and the genetic algorithm provides an attractive alternative to the classical techniques used to solve these problems.

Baffes, Paul↗