The theories of Dolan and Edwards [Proc. R. Soc. London Ser. A 337, 509 (1974)] and Eisenriegler ... more The theories of Dolan and Edwards [Proc. R. Soc. London Ser. A 337, 509 (1974)] and Eisenriegler et al. [J. Chem. Phys. 77, 6296 (1982)] for Gaussian chains confined between parallel plates and to a half-space are generalized to chains having arbitrary stiffness. The generalized theory exploits a recently discovered relation between semiflexible polymers and Euclidean-type Dirac fermions in which
We have used quasi-elastic neutron scattering techniques to study the dynamics of solvent water p... more We have used quasi-elastic neutron scattering techniques to study the dynamics of solvent water protons in H_ 2O solutions of deuterated poly(ethylene oxide) (Polyox) at room temperature. Using a slightly modified technique, we also have obtained values for the mean-square vibrational amplitude of polymer protons in powder and D_ 2O solutions of Polyox and trypsin at temperatures between 300K and
A study was conducted in November 1999 to assess sediment quality and condition of benthic fauna ... more A study was conducted in November 1999 to assess sediment quality and condition of benthic fauna in the Neuse River Estuary (NRE), North Carolina, USA, following the passage of three Atlantic hurricanes during the two months prior. Samples for analysis of macroinfauna (>0.5 mm sieve size), chemical contamination of sediments, and other abiotic environmental variables (salinity, dissolved oxygen, pH, depth, sediment granulometry) were collected at 20 sites from the mouth of the Neuse River at Pamlico Sound to approximately 90 km upstream. Results were compared to those obtained from the same area in July 1998 using similar protocols. Depressed salinity, caused by extreme rainfall and associated high freshwater flow, persisted throughout much of the estuary, which had experienced periods of water-column stratification and hypoxia of underlying waters. Fifteen of the 20 sites, representing 299 km 2 (76% of the survey area), also showed signs of benthic stress based on a multi-metric benthic index of biotic integrity (B-IBI). Benthic impacts included reductions in the abundance, diversity, and numbers of species and shifts in taxonomic composition, with a notable increase in dominance of the opportunistic polychaete Mediomastus ambiseta as other former dominant species declined. There was no significant increase in the extent of chemical contamination compared to pre-hurricane conditions. Storm-related reductions in dissoved oxygen and salinity were the more likely causes of the observed benthic impacts, though it was not possible, based on these results, to separate storm effects from seasonal changes in the benthos and annual episodes of summer anoxia and hypoxia.
Lombó, F., Künzel, E., Prado, L., Braña, AF, Bindseil, KU, Frevert, J., Bearden, D., Méndez, C., ... more Lombó, F., Künzel, E., Prado, L., Braña, AF, Bindseil, KU, Frevert, J., Bearden, D., Méndez, C., Salas, JA and Rohr, J.(2000), Die antitumorwirksame Hybrid-Verbindung Premithramycinon H verweist indirekt auf ein tricyclisches Intermediat der Biosynthese ...
Metabolomics : Official journal of the Metabolomic Society
Zebra mussel, Dreissena polymorpha, in the Great Lakes is being monitored as a bio-indicator orga... more Zebra mussel, Dreissena polymorpha, in the Great Lakes is being monitored as a bio-indicator organism for environmental health effects by the National Oceanic and Atmospheric Administration's Mussel Watch program. In order to monitor the environmental effects of industrial pollution on the ecosystem, invasive zebra mussels were collected from four stations-three inner harbor sites (LMMB4, LMMB1, and LMMB) in Milwaukee Estuary, and one reference site (LMMB5) in Lake Michigan, Wisconsin. Nuclear magnetic resonance (NMR)-based metabolomics was used to evaluate the metabolic profiles of the mussels from these four sites. The objective was to observe whether there were differences in metabolite profiles between impacted sites and the reference site; and if there were metabolic profile differences among the impacted sites. Principal component analyses indicated there was no significant difference between two impacted sites: north Milwaukee harbor (LMMB and LMMB4) and the LMMB5 referen...
The metabolomic fingerprints of a protected sea turtle species have been investigated for the fir... more The metabolomic fingerprints of a protected sea turtle species have been investigated for the first time using a nuclear magnetic resonance (NMR)-based metabolomics approach. We emphasized method development of optimal plasma filtration conditions (filter type, washing techniques, extract stability) for green turtles and other organisms, while also using the National Institute of Standards and Technology (NIST) Standard Reference Material 1950 (SRM 1950) Metabolites in Human Plasma for quality control. We surveyed the blood plasma metabolomic fingerprints of Hawaiian green sea turtles representing a wide range of physiological conditions that include varying disease states, behavioral conditions, and locales. The turtles sampled were free-swimming (n=5 from Hualalai on the west coast of the island of Hawaii), basking (n=7 from Hualalai), free-swimming tumor-free (n=3 from Kapoho, east coast of Hawaii), or freeswimming afflicted with external tumors (n=5 from Kapoho) caused by the disease, fibropapillomatosis (FP). The metabolomic profiles and the specific metabolites that differed among individual turtles are discussed. This optimized tool and the annotated metabolic profiles will benefit future investigations into the behavioral and disease conditions of the green turtle.
Coral bleaching occurs when the symbioses between coral animals and their zooxanthellae is 27 dis... more Coral bleaching occurs when the symbioses between coral animals and their zooxanthellae is 27 disrupted, either as part of a natural cycle or as the result of unusual events. The bacterium 28
The objective of this study was to determine inorganic and organic contaminant concentrations in ... more The objective of this study was to determine inorganic and organic contaminant concentrations in edible tissue of fish collected from eight coastal areas receiving wastewater discharges and from two reference locations. Trace metal residues were statistically similar regardless of the collection site. Zinc (100% detection in all samples), total mercury (100%), total arsenic (92%), copper (92%), and selenium (88%) were the more commonly detected trace metals. Mercury concentrations exceeded the Florida health-based standard of 0.5 microg/g for limited fish consumption in 30% of the total samples and averaged 0.40 (+/- 1 S.D. = 0.22, range < or = 0.08 to 0.85) microg/g wet weight. The average total PAH concentrations were 1.79 (+/- 1.60) ng/g (reference areas) and 2.17 (+/- 3.29) ng/g (wastewater-impacted areas). Pyrene was detected most frequently (63% of the total samples) and averaged 0.74 (+/- 0.35) ng/g wet wt. The average total PCB concentrations were 4.8 (+/- 7.1) ng/g (reference areas) and 31.6 (+/- 31.3) ng/g (wastewater-impacted areas) Concentrations of dieldrin and cis-chlordane were approximately eight times greater, respectively, in fish collected from wastewater receiving waters, whereas total DDT and total pesticide concentrations were not elevated in the same areas. Concentrations of total PCBs and all chlorinated pesticides were below US health-based standards. The lack of a published reference data base for fish tissue quality in near-coastal areas of the Gulf of Mexico restricts an assessment of the environmental significance of results from this and similar studies investigating the fate of point source contaminants.
b-glucan is a (1R3)-b-linked glucose polymer with (1R6)-b-linked side chains and a major componen... more b-glucan is a (1R3)-b-linked glucose polymer with (1R6)-b-linked side chains and a major component of fungal cell walls. b-glucans provide structural integrity to the fungal cell wall. The nature of the (1-6)-b-linked side chain structure of fungal (1R3,1R6)-b-D-glucans has been very difficult to elucidate. Herein, we report the first detailed structural characterization of the (1R6)-b-linked side chains of Candida glabrata using high-field NMR. The (1R6)-b-linked side chains have an average length of 4 to 5 repeat units spaced every 21 repeat units along the (1R3)-linked polymer backbone. Computer modeling suggests that the side chains have a bent curve structure that allows for a flexible interconnection with parallel (1R3)-b-Dglucan polymers, and/or as a point of attachment for proteins. Based on these observations we propose new approaches to how (1R6)-b-linked side chains interconnect with neighboring glucan polymers in a manner that maximizes fungal cell wall strength, while also allowing for flexibility, or plasticity.
With the increasing use of metabolomics as a means to study a large number of different biologica... more With the increasing use of metabolomics as a means to study a large number of different biological research questions, there is a need for a minimal set of reporting standards that allow the scientific community to evaluate, understand, repeat, compare and re-investigate metabolomics studies. Here we propose, a first draft of minimal requirements to effectively describe the biological context of metabolomics studies that involve microbial or in vitro biological subjects. This recommendation has been produced by the microbiology and in vitro biology working subgroup of the Metabolomics Standards Initiative in collaboration with the yeast systems biology network as part of a wider standardization initiative led by the Metabolomics Society. Microbial and in vitro biology metabolomics is defined by this sub-working group as studies with any cell or organism that require a defined external medium to facilitate growth and propagation. Both a minimal set and a best practice set of reporting standards for metabolomics experiments have been defined. The minimal set of reporting standards for microbial or in vitro biology metabolomics experiments includes those factors that are specific for metabolomics experiments and that critically determine the outcome of the experiments. The best practice set of reporting standards contains both the factors that
Mithramycin is an aureolic acid-type antimicrobial and antitumor agent produced by Streptomyces a... more Mithramycin is an aureolic acid-type antimicrobial and antitumor agent produced by Streptomyces argillaceus. Modifying post-polyketide synthase (PKS) tailoring enzymes involved in the production of mithramycin is an effective way of gaining further information regarding the late steps of its biosynthetic pathway. In addition, new "unnatural" natural products of the aureolic acid-type class are likely to be produced. The role of two such post-PKS tailoring enzymes, encoded by mtmC and mtmTIII, was investigated, and four novel aureolic acid class drugs, two premithramycin-type molecules and two mithramycin derivatives, were isolated from mutant strains constructed by insertional gene inactivation of either of these two genes. From data bank comparisons, the corresponding proteins MtmC and MtmTIII were believed to act as a C-methyltransferase involved in the production of the D-mycarose (sugar E) of mithramycin and as a ketoreductase seemingly involved in the biosynthesis of the mithramycin aglycon, respectively. However, gene inactivation and analysis of the accumulated products revealed that both genes encode enzymes participating in the biosynthesis of the D-mycarose building block. Furthermore, the inactivation of MtmC seems to affect the ketoreductase responsible for 4-ketoreduction of sugar C, a D-olivose. Instead of obtaining premithramycin and mithramycin derivatives with a modified E-sugar upon inactivation of mtmC, compounds were obtained that completely lack the E-sugar moiety and that possess an unexpected 4-ketosugar moiety instead of the D-olivose at the beginning of the lower deoxysaccharide chain. The inactivation of mtmTIII led to the accumulation of 4E-ketomithramycin, showing that this ketoreductase is responsible for the 4-ketoreduction of the D-mycarose moiety. The new compounds of the mutant strains, 4A-ketopremithramycin A2, 4A-keto-9-demethylpremithramycin A2, 4C-keto-demycarosylmithramycin, and 4E-ketomithramycin, indicate surprising substrate flexibility of post-PKS enzymes of the mithramycin biosynthetic pathway. Although the glycosyltransferase responsible for the attachment of D-mycarose cannot transfer the unmethylated sugar to the existing lower disaccharide chain, it can transfer the 4-ketoform of sugar E. In addition, the glycosyltransferase MtmGIV, which is responsible for the linkage of sugar C, is also able to transfer an activated 4-ketosugar. The oxygenase MtmOIV, normally responsible for the oxidative cleavage of the tetracyclic premithramycin B into the tricyclic immediate precursor of mithramycin, can act on a substrate analogue with a modified or even incomplete trisaccharide chain. The same is true for glycosyltransferases MtmGI and MtmGII, both of which partake in the formation and attachment of the A-B disaccharide in mithramycin.
Our investigations on the discovery of novel natural metabolites using type II polyketide synthas... more Our investigations on the discovery of novel natural metabolites using type II polyketide synthase gene probes (actI/III) yielded an unusual angucyclinone, oviedomycin , when applied to the oleandomycin (1) producer Streptomyces antibioticus ATCC11891. The novel natural product was produced using S. albus R -Mas a host strain, into which a cosmid containing the oviedomycin gene cluster was transformed. Its structure was elucidated by NMR spectroscopy and mass spectrometry.
The theories of Dolan and Edwards [Proc. R. Soc. London Ser. A 337, 509 (1974)] and Eisenriegler ... more The theories of Dolan and Edwards [Proc. R. Soc. London Ser. A 337, 509 (1974)] and Eisenriegler et al. [J. Chem. Phys. 77, 6296 (1982)] for Gaussian chains confined between parallel plates and to a half-space are generalized to chains having arbitrary stiffness. The generalized theory exploits a recently discovered relation between semiflexible polymers and Euclidean-type Dirac fermions in which
We have used quasi-elastic neutron scattering techniques to study the dynamics of solvent water p... more We have used quasi-elastic neutron scattering techniques to study the dynamics of solvent water protons in H_ 2O solutions of deuterated poly(ethylene oxide) (Polyox) at room temperature. Using a slightly modified technique, we also have obtained values for the mean-square vibrational amplitude of polymer protons in powder and D_ 2O solutions of Polyox and trypsin at temperatures between 300K and
A study was conducted in November 1999 to assess sediment quality and condition of benthic fauna ... more A study was conducted in November 1999 to assess sediment quality and condition of benthic fauna in the Neuse River Estuary (NRE), North Carolina, USA, following the passage of three Atlantic hurricanes during the two months prior. Samples for analysis of macroinfauna (>0.5 mm sieve size), chemical contamination of sediments, and other abiotic environmental variables (salinity, dissolved oxygen, pH, depth, sediment granulometry) were collected at 20 sites from the mouth of the Neuse River at Pamlico Sound to approximately 90 km upstream. Results were compared to those obtained from the same area in July 1998 using similar protocols. Depressed salinity, caused by extreme rainfall and associated high freshwater flow, persisted throughout much of the estuary, which had experienced periods of water-column stratification and hypoxia of underlying waters. Fifteen of the 20 sites, representing 299 km 2 (76% of the survey area), also showed signs of benthic stress based on a multi-metric benthic index of biotic integrity (B-IBI). Benthic impacts included reductions in the abundance, diversity, and numbers of species and shifts in taxonomic composition, with a notable increase in dominance of the opportunistic polychaete Mediomastus ambiseta as other former dominant species declined. There was no significant increase in the extent of chemical contamination compared to pre-hurricane conditions. Storm-related reductions in dissoved oxygen and salinity were the more likely causes of the observed benthic impacts, though it was not possible, based on these results, to separate storm effects from seasonal changes in the benthos and annual episodes of summer anoxia and hypoxia.
Lombó, F., Künzel, E., Prado, L., Braña, AF, Bindseil, KU, Frevert, J., Bearden, D., Méndez, C., ... more Lombó, F., Künzel, E., Prado, L., Braña, AF, Bindseil, KU, Frevert, J., Bearden, D., Méndez, C., Salas, JA and Rohr, J.(2000), Die antitumorwirksame Hybrid-Verbindung Premithramycinon H verweist indirekt auf ein tricyclisches Intermediat der Biosynthese ...
Metabolomics : Official journal of the Metabolomic Society
Zebra mussel, Dreissena polymorpha, in the Great Lakes is being monitored as a bio-indicator orga... more Zebra mussel, Dreissena polymorpha, in the Great Lakes is being monitored as a bio-indicator organism for environmental health effects by the National Oceanic and Atmospheric Administration's Mussel Watch program. In order to monitor the environmental effects of industrial pollution on the ecosystem, invasive zebra mussels were collected from four stations-three inner harbor sites (LMMB4, LMMB1, and LMMB) in Milwaukee Estuary, and one reference site (LMMB5) in Lake Michigan, Wisconsin. Nuclear magnetic resonance (NMR)-based metabolomics was used to evaluate the metabolic profiles of the mussels from these four sites. The objective was to observe whether there were differences in metabolite profiles between impacted sites and the reference site; and if there were metabolic profile differences among the impacted sites. Principal component analyses indicated there was no significant difference between two impacted sites: north Milwaukee harbor (LMMB and LMMB4) and the LMMB5 referen...
The metabolomic fingerprints of a protected sea turtle species have been investigated for the fir... more The metabolomic fingerprints of a protected sea turtle species have been investigated for the first time using a nuclear magnetic resonance (NMR)-based metabolomics approach. We emphasized method development of optimal plasma filtration conditions (filter type, washing techniques, extract stability) for green turtles and other organisms, while also using the National Institute of Standards and Technology (NIST) Standard Reference Material 1950 (SRM 1950) Metabolites in Human Plasma for quality control. We surveyed the blood plasma metabolomic fingerprints of Hawaiian green sea turtles representing a wide range of physiological conditions that include varying disease states, behavioral conditions, and locales. The turtles sampled were free-swimming (n=5 from Hualalai on the west coast of the island of Hawaii), basking (n=7 from Hualalai), free-swimming tumor-free (n=3 from Kapoho, east coast of Hawaii), or freeswimming afflicted with external tumors (n=5 from Kapoho) caused by the disease, fibropapillomatosis (FP). The metabolomic profiles and the specific metabolites that differed among individual turtles are discussed. This optimized tool and the annotated metabolic profiles will benefit future investigations into the behavioral and disease conditions of the green turtle.
Coral bleaching occurs when the symbioses between coral animals and their zooxanthellae is 27 dis... more Coral bleaching occurs when the symbioses between coral animals and their zooxanthellae is 27 disrupted, either as part of a natural cycle or as the result of unusual events. The bacterium 28
The objective of this study was to determine inorganic and organic contaminant concentrations in ... more The objective of this study was to determine inorganic and organic contaminant concentrations in edible tissue of fish collected from eight coastal areas receiving wastewater discharges and from two reference locations. Trace metal residues were statistically similar regardless of the collection site. Zinc (100% detection in all samples), total mercury (100%), total arsenic (92%), copper (92%), and selenium (88%) were the more commonly detected trace metals. Mercury concentrations exceeded the Florida health-based standard of 0.5 microg/g for limited fish consumption in 30% of the total samples and averaged 0.40 (+/- 1 S.D. = 0.22, range < or = 0.08 to 0.85) microg/g wet weight. The average total PAH concentrations were 1.79 (+/- 1.60) ng/g (reference areas) and 2.17 (+/- 3.29) ng/g (wastewater-impacted areas). Pyrene was detected most frequently (63% of the total samples) and averaged 0.74 (+/- 0.35) ng/g wet wt. The average total PCB concentrations were 4.8 (+/- 7.1) ng/g (reference areas) and 31.6 (+/- 31.3) ng/g (wastewater-impacted areas) Concentrations of dieldrin and cis-chlordane were approximately eight times greater, respectively, in fish collected from wastewater receiving waters, whereas total DDT and total pesticide concentrations were not elevated in the same areas. Concentrations of total PCBs and all chlorinated pesticides were below US health-based standards. The lack of a published reference data base for fish tissue quality in near-coastal areas of the Gulf of Mexico restricts an assessment of the environmental significance of results from this and similar studies investigating the fate of point source contaminants.
b-glucan is a (1R3)-b-linked glucose polymer with (1R6)-b-linked side chains and a major componen... more b-glucan is a (1R3)-b-linked glucose polymer with (1R6)-b-linked side chains and a major component of fungal cell walls. b-glucans provide structural integrity to the fungal cell wall. The nature of the (1-6)-b-linked side chain structure of fungal (1R3,1R6)-b-D-glucans has been very difficult to elucidate. Herein, we report the first detailed structural characterization of the (1R6)-b-linked side chains of Candida glabrata using high-field NMR. The (1R6)-b-linked side chains have an average length of 4 to 5 repeat units spaced every 21 repeat units along the (1R3)-linked polymer backbone. Computer modeling suggests that the side chains have a bent curve structure that allows for a flexible interconnection with parallel (1R3)-b-Dglucan polymers, and/or as a point of attachment for proteins. Based on these observations we propose new approaches to how (1R6)-b-linked side chains interconnect with neighboring glucan polymers in a manner that maximizes fungal cell wall strength, while also allowing for flexibility, or plasticity.
With the increasing use of metabolomics as a means to study a large number of different biologica... more With the increasing use of metabolomics as a means to study a large number of different biological research questions, there is a need for a minimal set of reporting standards that allow the scientific community to evaluate, understand, repeat, compare and re-investigate metabolomics studies. Here we propose, a first draft of minimal requirements to effectively describe the biological context of metabolomics studies that involve microbial or in vitro biological subjects. This recommendation has been produced by the microbiology and in vitro biology working subgroup of the Metabolomics Standards Initiative in collaboration with the yeast systems biology network as part of a wider standardization initiative led by the Metabolomics Society. Microbial and in vitro biology metabolomics is defined by this sub-working group as studies with any cell or organism that require a defined external medium to facilitate growth and propagation. Both a minimal set and a best practice set of reporting standards for metabolomics experiments have been defined. The minimal set of reporting standards for microbial or in vitro biology metabolomics experiments includes those factors that are specific for metabolomics experiments and that critically determine the outcome of the experiments. The best practice set of reporting standards contains both the factors that
Mithramycin is an aureolic acid-type antimicrobial and antitumor agent produced by Streptomyces a... more Mithramycin is an aureolic acid-type antimicrobial and antitumor agent produced by Streptomyces argillaceus. Modifying post-polyketide synthase (PKS) tailoring enzymes involved in the production of mithramycin is an effective way of gaining further information regarding the late steps of its biosynthetic pathway. In addition, new "unnatural" natural products of the aureolic acid-type class are likely to be produced. The role of two such post-PKS tailoring enzymes, encoded by mtmC and mtmTIII, was investigated, and four novel aureolic acid class drugs, two premithramycin-type molecules and two mithramycin derivatives, were isolated from mutant strains constructed by insertional gene inactivation of either of these two genes. From data bank comparisons, the corresponding proteins MtmC and MtmTIII were believed to act as a C-methyltransferase involved in the production of the D-mycarose (sugar E) of mithramycin and as a ketoreductase seemingly involved in the biosynthesis of the mithramycin aglycon, respectively. However, gene inactivation and analysis of the accumulated products revealed that both genes encode enzymes participating in the biosynthesis of the D-mycarose building block. Furthermore, the inactivation of MtmC seems to affect the ketoreductase responsible for 4-ketoreduction of sugar C, a D-olivose. Instead of obtaining premithramycin and mithramycin derivatives with a modified E-sugar upon inactivation of mtmC, compounds were obtained that completely lack the E-sugar moiety and that possess an unexpected 4-ketosugar moiety instead of the D-olivose at the beginning of the lower deoxysaccharide chain. The inactivation of mtmTIII led to the accumulation of 4E-ketomithramycin, showing that this ketoreductase is responsible for the 4-ketoreduction of the D-mycarose moiety. The new compounds of the mutant strains, 4A-ketopremithramycin A2, 4A-keto-9-demethylpremithramycin A2, 4C-keto-demycarosylmithramycin, and 4E-ketomithramycin, indicate surprising substrate flexibility of post-PKS enzymes of the mithramycin biosynthetic pathway. Although the glycosyltransferase responsible for the attachment of D-mycarose cannot transfer the unmethylated sugar to the existing lower disaccharide chain, it can transfer the 4-ketoform of sugar E. In addition, the glycosyltransferase MtmGIV, which is responsible for the linkage of sugar C, is also able to transfer an activated 4-ketosugar. The oxygenase MtmOIV, normally responsible for the oxidative cleavage of the tetracyclic premithramycin B into the tricyclic immediate precursor of mithramycin, can act on a substrate analogue with a modified or even incomplete trisaccharide chain. The same is true for glycosyltransferases MtmGI and MtmGII, both of which partake in the formation and attachment of the A-B disaccharide in mithramycin.
Our investigations on the discovery of novel natural metabolites using type II polyketide synthas... more Our investigations on the discovery of novel natural metabolites using type II polyketide synthase gene probes (actI/III) yielded an unusual angucyclinone, oviedomycin , when applied to the oleandomycin (1) producer Streptomyces antibioticus ATCC11891. The novel natural product was produced using S. albus R -Mas a host strain, into which a cosmid containing the oviedomycin gene cluster was transformed. Its structure was elucidated by NMR spectroscopy and mass spectrometry.
Metabolomic technologies are increasingly being applied to study biological questions in a range ... more Metabolomic technologies are increasingly being applied to study biological questions in a range of different settings from clinical through to environmental. As with other high-throughput technologies, such as those used in transcriptomics and proteomics, metabolomics continues to generate large volumes of complex data that necessitates computational management. Making sense of this wealth of information also requires access to sufficiently detailed and well annotated meta-data. Here we provide standard reporting requirements for describing biological samples, taken from an environmental context and involved in metabolomic experiments. It is our intention that these reporting requirements should guide and support the standardised annotation, dissemination
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Papers by Daniel Bearden