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J P Allen

Publications and source records attributed to J P Allen.

At least 37 records · Page 2Linked to original sources

Proton release upon oxidation of tyrosine in reaction centers from Rhodobacter sphaeroides.

Markedly different light-induced protonational changes were measured in two reaction center mutants of Rhodobacter sphaeroides. A quadruple mutant containing alterations, at residues L131, M160, M197, and M210, that elevate the midpoint potential of the bacteriochlorophyll dimer was compared to the Y(M) mutant, which contains these alterations plus a tyrosine at M164 serving as a secondary electron donor [Kálmán et al., Nature 402 (1999) 696]. In the quadruple mutant, a proton uptake of 0.1-0.3 H(+)/reaction center between pH 6 and 10 resulted from formation of the oxidized bacteriochlorophyll donor and reduced primary quinone. In the Y(M) mutant, a maximal proton release of -0.5 H(+)/reaction center at pH 8 was attributed to formation of the tyrosyl radical and modeled using electrostatic and direct proton-releasing mechanisms.

Amino Acid Substitution↗

The legacy of Biosphere 2 for the study of biospherics and closed ecological systems.

The unprecedented challenges of creating Biosphere 2, the world's first laboratory for biospherics, the study of global ecology and long-term closed ecological system dynamics, led to breakthrough developments in many fields, and a deeper understanding of the opportunities and difficulties of material closure. This paper will review accomplishments and challenges, citing some of the key research findings and publications that have resulted from the experiments in Biosphere 2. Engineering accomplishments included development of a technique for variable volume to deal with pressure differences between the facility and outside environment, developing methods of atmospheric leak detection and sealing, while achieving new standards of closure, with an annual atmospheric leakrate of less than 10%, or less than 300 ppm per day. This degree of closure permitted detailed tracking of carbon dioxide, oxygen, and trace gases such as nitrous oxide and ethylene over the seasonal variability of two years. Full closure also necessitated developing new approaches and technologies for complete air, water, and wastewater recycle and reuse within the facility. The development of a soil-based highly productive agricultural system was a first in closed ecological systems, and much was learned about managing a wide variety of crops using non-chemical means of pest and disease control. Closed ecological systems have different temporal biogeochemical cycling and ranges of atmospheric components because of their smaller reservoirs of air, water and soil, and higher concentration of biomass, and Biosphere 2 provided detailed examination and modeling of these accelerated cycles over a period of closure which measured in years. Medical research inside Biosphere 2 included the effects on humans of lowered oxygen: the discovery that human productivity can be maintained with good health with lowered atmospheric oxygen levels could lead to major economies on the design of space stations and planetary/lunar settlements. The improved health resulting from the calorie-restricted but nutrient dense Biosphere 2 diet was the first such scientifically controlled experiment with humans. The success of Biosphere 2 in creating a diversity of terrestrial and marine environments, from rainforest to coral reef, allowed detailed studies with comprehensive measurements such that the dynamics of these complex biomic systems are now better understood. The coral reef ecosystem, the largest artificial reef ever built, catalyzed methods of study now being applied to planetary coral reef systems. Restoration ecology advanced through the creation and study of the dynamics of adaptation and self-organization of the biomes in Biosphere 2. The international interest that Biosphere 2 generated has given new impetus to the public recognition of the sciences of biospheres (biospherics), biomes and closed ecological life systems. The facility, although no longer a materially-closed ecological system, is being used as an educational facility by Columbia University as an introduction to the study of the biosphere and complex system ecology and for carbon dioxide impacts utilizing the complex ecosystems created in Biosphere '. The many lessons learned from Biosphere 2 are being used by its key team of creators in their design and operation of a laboratory-sized closed ecological system, the Laboratory Biosphere, in operation as of March 2002, and for the design of a Mars on Earth(TM) prototype life support system for manned missions to Mars and Mars surface habitats. Biosphere 2 is an important foundation for future advances in biospherics and closed ecological system research.

Agriculture↗

Initial experimental results from the Laboratory Biosphere closed ecological system facility.

An initial experiment in the Laboratory Biosphere facility, Santa Fe, New Mexico, was conducted May-August 2002 using a soil-based system with light levels (at 12 h per day) of 58-mol m-2 d-1. The crop tested was soybean, cultivar Hoyt, which produced an aboveground biomass of 2510 grams. Dynamics of a number of trace gases showed that methane, nitrous oxide, carbon monoxide, and hydrogen gas had initial increases that were substantially reduced in concentration by the end of the experiment. Methane was reduced from 209 ppm to 11 ppm, and nitrous oxide from 5 ppm to 1.4 ppm in the last 40 days of the closure experiment. Ethylene was at elevated levels compared to ambient during the flowering/fruiting phase of the crop. Soil respiration from the 5.37 m2 (1.46 m3) soil component was estimated at 23.4 ppm h-1 or 1.28 g CO2 h-1 or 5.7 g CO2 m-2 d-1. Phytorespiration peaked near the time of fruiting at about 160 ppm h-1. At the height of plant growth, photosynthesis CO2 draw down was as high as 3950 ppm d-1, and averaged 265 ppm h-1 (whole day averages) during lighted hours with a range of 156-390 ppm h-1. During this period, the chamber required injections of CO2 to continue plant growth. Oxygen levels rose along with the injections of carbon dioxide. Upon several occasions, CO2 was allowed to be drawn down to severely limiting levels, bottoming at around 150 ppm. A strong positive correlation (about 0.05 ppm h-1 ppm-1 with r2 about 0.9 for the range 1000-5000 ppm) was observed between atmospheric CO2 concentration and the rate of fixation up to concentrations of around 8800 ppm CO2.

Biomass↗

Influence of the protein environment on the properties of a tyrosyl radical in reaction centers from Rhodobacter sphaeroides.

The influence of the local environment on the formation of a tyrosyl radical was investigated in modified photosynthetic reaction centers from Rhodobacter sphaeroides. The reaction centers contain a tyrosine residue placed approximately 10 A from a highly oxidizing bacteriochlorophyll dimer. Measurements by both optical and electron paramagnetic resonance spectroscopy revealed spectral features that are assigned as arising primarily from an oxidized bacteriochlorophyll dimer at low pH values and from a tyrosyl radical at high pH values, with a well-defined transition that occurred with a pK(a) of 6.9. A model based on the wild-type structure indicated that the Tyr at M164 is likely to form a hydrogen bond with His M193 and to interact weakly with Glu M173. Substitution of Tyr or Glu for His at M193 increased the pK(a) for the transition from 6.9 to 8.9, while substitution of Gln for His M193 resulted in a higher pK(a) value. Substitution of Glu M173 with Gln resulted in loss of the partial formation of the tyrosyl that occurs in the other mutants at low pH values. The results are interpreted in terms of the ability of the residues to act as proton acceptors for the oxidized tyrosine, with the pK(a) values reflecting those of either the putative proton acceptor or the tyrosine, in accord with general models of amino acid radicals.

Arginine↗

The influence of detergents on the solubility of membrane proteins.

The relationship between the effect of detergents and amphiphiles on protein solubility and their use in crystallization solutions was examined for the reaction center from Rhodobacter sphaeroides. Measurement by a centrifugation assay of the solubility of the reaction center as a function of ionic strength revealed dramatic differences in the intrinsic solubility at zero ionic strength in the presence of various detergents and amphiphiles. High protein-solubility values were found for beta-octyl glucoside and for lauryldimethylamine-N-oxide with heptanetriol. The solubility differences are interpreted in terms of fundamental properties such as the polarity of the detergent molecules. Conditions that resulted in high protein solubility correspond to conditions that have been shown to be successful for crystallization of the reaction center. These results suggest that crystallization is favored for detergents and amphiphiles that optimize the solubility of integral membrane proteins.

Detergents↗

Interactions between lipids and bacterial reaction centers determined by protein crystallography.

The structure of the reaction center from Rhodobacter sphaeroides has been solved by using x-ray diffraction at a 2.55-A resolution limit. Three lipid molecules that lie on the surface of the protein are resolved in the electron density maps. In addition to a cardiolipin that has previously been reported [McAuley, K. E., Fyfe, P. K., Ridge, J. P., Isaacs, N. W., Cogdell, R. J. & Jones, M. R. (1999) Proc. Natl. Acad. Sci. USA 96, 14706-14711], two other major lipids of the cell membrane are found, a phosphatidylcholine and a glucosylgalactosyl diacylglycerol. The presence of these three lipids has been confirmed by laser mass spectroscopy. The lipids are located in the hydrophobic region of the protein surface and interact predominately with hydrophobic amino acids, in particular aromatic residues. Although the cardiolipin is over 15 A from the cofactors, the other two lipids are in close contact with the cofactors and may contribute to the difference in energetics for the two branches of cofactors that is primarily responsible for the asymmetry of electron transfer. The glycolipid is 3.5 A from the active bacteriochlorophyll monomer and shields this cofactor from the solvent in contrast to a much greater exposed surface evident for the inactive bacteriochlorophyll monomer. The phosphate atom of phosphatidylcholine is 6.5 A from the inactive bacteriopheophytin, and the associated electrostatic interactions may contribute to electron transfer rates involving this cofactor. Overall, the lipids span a distance of approximately 30 A, which is consistent with a bilayer-like arrangement suggesting the presence of an "inner shell" of lipids around membrane proteins that is critical for membrane function.

Cardiolipins↗

The structure of the heterodimer reaction center from Rhodobacter sphaeroides at 2.55 å resolution.

Crystals have been obtained of reaction centers of the heterodimer mutant that has significantly different properties than wild type due to the primary donor being formed from both a bacteriochlorophyll and bacteriopheophytin rather than two bacteriochlorophylls as found for wild type. The crystals belong to the trigonal space group P3(1)21 and the structure has been refined to a resolution limit of 2.55 A with an R factor of 19.0%. The electron density maps confirm that a primary donor does indeed contain a bacteriopheophytin due to the His to Leu substitution at M202 that coordinates the corresponding bacteriochlorophyll in wild-type. Other structural changes compared to wild type are relatively minor with the relative orientation and positioning of the two tetrapyrroles forming the primary donor being unchanged within the error. Compared to wild type, the only significant alterations are small shifts of residues M196 to M206, a rotation of the side chain of Ile M206, and the loss of a bound water molecule that in wild-type is hydrogen-bonded to both His M202 and the bacteriochlorophyll monomer on the active branch. Since hydrogen-bonding interactions strongly influence the energies of tetrapyrroles, the loss of the water molecule should result in changes in the energies of the bacteriochlorophyll monomer that contributes to the observed functional differences with wild-type.

Journal Article↗

Potential integration of wetland wastewater treatment with space life support systems.

Subsurface-flow constructed wetlands for wastewater treatment and nutrient recycling have a number of advantages in planetary exploration scenarios: they are odorless, relatively low labor and low energy, assist in purification of water and recycling of atmospheric CO2, and can directly grow some food crops. This article presents calculations for integration of wetland wastewater treatment with a prototype ground-based experimental facility ("Mars on Earth") supporting four people showing that an area of 4-6 m2 may be sufficient to accomplish wastewater treatment and recycling. Discharge water from the wetland system can be used as irrigation water for the agricultural crop area, thus ensuring complete reclamation and utilization of nutrients within the bioregenerative life support system. Because the primary requirements for wetland treatment systems are warm temperatures and lighting, such bioregenerative systems can be integrated into space life support systems because heat from the lights may be used for temperature maintenance in the human living environment. Subsurface-flow wetlands can be modified for space habitats to lower space and mass requirements. Many of its construction requirements can eventually be met with use of in situ materials, such as gravel from the Mars surface. Because the technology does not depend on machinery and chemicals, and relies more on natural ecological mechanisms (microbial and plant metabolism), maintenance requirements (e.g., pumps, aerators, and chemicals) are minimized, and systems may have long operating lifetimes. Research needs include suitability of Martian soil and gravel for wetland systems, system sealing and liner options in a Mars base, and determination of wetland water quality efficiency under varying temperature and light regimes.

Ecological Systems, Closed↗

Light, plants, and power for life support on Mars.

Regardless of how well other growing conditions are optimized, crop yields will be limited by the available light up to saturation irradiances. Considering the various factors of clouds on Earth, dust storms on Mars, thickness of atmosphere, and relative orbits, there is roughly 2/3 as much light averaged annually on Mars as on Earth. On Mars, however, crops must be grown under controlled conditions (greenhouse or growth rooms). Because there presently exists no material that can safely be pressurized, insulated, and resist hazards of puncture and deterioration to create life support systems on Mars while allowing for sufficient natural light penetration as well, artificial light will have to be supplied. If high irradiance is provided for long daily photoperiods, the growing area can be reduced by a factor of 3-4 relative to the most efficient irradiance for cereal crops such as wheat and rice, and perhaps for some other crops. Only a small penalty in required energy will be incurred by such optimization. To obtain maximum yields, crops must be chosen that can utilize high irradiances. Factors that increase ability to convert high light into increased productivity include canopy architecture, high-yield index (harvest index), and long-day or day-neutral flowering and tuberization responses. Prototype life support systems such as Bios-3 in Siberia or the Mars on Earth Project need to be undertaken to test and further refine systems and parameters.

Biomass↗

Electrostatic interactions between charged amino acid residues and the bacteriochlorophyll dimer in reaction centers from Rhodobacter sphaeroides.

The extent of electrostatic contributions from the protein environment was assessed by the introduction of ionizable residues near the bacteriochlorophyll dimer in reaction centers from Rhodobacter sphaeroides. Two mutations at symmetry-related sites, M199 Asn to Asp and L170 Asn to Asp, resulted in a 48 and 44 mV lowering of the midpoint potential, respectively, compared to the wild type at pH 8, while a 75 mV decrease in the midpoint potential was observed for the mutation L168 His to Glu. The decrease relative to wild type was found to be approximately additive, up to 147 mV, for various combinations of the mutations. As the pH was lowered from 9.5 to 6.0, the relative decrease in the midpoint potential became smaller for each of these three mutations. Titration of the pH dependence of the change in midpoint potential of the M199 Asn to Asp mutant compared to wild type yielded a pK(a) value of 7.9 and a change in midpoint potential from low to high pH of 59 mV. The major effect of the mutation on the midpoint potential of the dimer is interpreted as stemming from a negative charge on the residue. An average dielectric constant of approximately 20 was estimated for the local protein environment, consistent with a relatively hydrophobic environment for residue M199. The rate of charge recombination between the primary quinone acceptor and the bacteriochlorophyll dimer decreased in the M199 Asn to Asp mutant at high pH, reflecting the decrease in midpoint potential.

Bacteriochlorophylls↗

Structure of cytochrome c2 from Rhodospirillum centenum.

Cytochrome c(2) from the purple photosynthetic bacterium Rhodospirillum centenum has been crystallized by the sitting-drop vapour-diffusion method. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 29.7, b = 59.9, c = 65.4 A, and diffract to a resolution limit of 1.7 A. The Fe-atom position was determined from its anomalous scattering contribution and a molecular-replacement solution was calculated. The correctness of the solution was confirmed by parallel isomorphous replacement studies. The resulting model has a type I cytochrome fold with two features, an extended alpha-helix and a surface-charge distribution, that are distinctive to this protein. The implications of these structural features for the ability of the cytochrome to serve as an electron carrier are discussed.

Crystallization↗

Individual interactions influence the crystalline order for membrane proteins.

The role of contact interactions in the crystallization of membrane proteins was assessed by mutation of amino-acid residues on the surface of the reaction center from Rhodobacter sphaeroides. Five single-site mutants were constructed, with changes in contact regions found in the trigonal and tetragonal forms but not the orthorhombic form. Crystallization trials for the tetragonal form yielded either no crystals or crystals with an altered morphology, whereas crystals grew in the other two forms, indicating that these interactions are essential for the stability of the tetragonal crystals. Changes in the structures determined by X-ray diffraction of trigonal crystals for each mutant were related to the quality of the diffraction. Significant differences in the resolution limit of the crystals were associated with the loss of specific interactions between neighboring proteins. The results suggest that the contact regions are crucial for obtaining highly ordered crystals of membrane proteins.

Bacterial Proteins↗

Amphiphiles modify the properties of detergent solutions used in crystallization of membrane proteins.

The effect of the amphiphile heptanetriol on the properties of solutions containing several detergents commonly used for crystallization of membrane proteins was characterized. The critical micelle concentration was found to be relatively unchanged by the presence of the amphiphile. In contrast, the addition of heptanetriol to solutions containing both detergent and polyethylene glycol exhibited significant shifts in the clouding behavior, with the largest shifts being for lauryl dimethylamine oxide. These results suggest that conditions favorable for crystallization of integral membrane proteins can be inferred from the properties of the detergents and amphiphiles.

Crystallization↗

The alcohol use disorders identification test: an aid to recognition of alcohol problems in primary care patients.

BACKGROUND: Misuse of alcohol is associated with a range of medical problems. Fortunately, a simple pencil-and-paper measure, the Alcohol Use Disorders Identification Test, can effectively and efficiently screen for early-stage alcohol abuse as well as provide the physician information that can assist in brief intervention. OBJECTIVE: The objective of this article is to briefly summarize research published on the Alcohol Use Disorders Identification Test and suggest its potential role in brief intervention in primary care settings. METHODS: Scientific literature on the Alcohol Use Disorders Identification Test though 2000 was reviewed and synthesized to address issues relevant to use of the test in primary health care settings. RESULTS: The Alcohol Use Disorders Identification Test is quite sensitive and specific and compares favorably with alternative self-report screens for alcohol problems.

Aged↗

The role of laboratory tests in alcoholism treatment.

Although assessment in the field of alcoholism treatment is generally verbal in nature, biological tests can also provide counselors and program evaluators useful and unique information. Five such laboratory measures are briefly described, with particular emphasis on carbohydrate deficient transferrin, a biomarker recently approved by the FDA. Applications for laboratory tests in alcohol screening, motivating patients, and monitoring treatment progress are also proposed.

Alanine Transaminase↗

Carbohydrate-deficient transferrin: an aid to early recognition of alcohol relapse.

Although the primary use of biochemical markers of heavy drinking is to assist in screening for alcohol problems, laboratory tests may also aid in early identification of relapse. This report reviews research findings on a new marker, carbohydrate deficient transferrin (CDT), in alcoholics receiving treatment or in follow-up. It also offers recommendations on how CDT may be employed by clinicians monitoring drinking status.

Alcoholism↗

Possible reasons why heavy drinking increases carbohydrate-deficient transferrin.

BACKGROUND: Transferrin is a globular protein synthesized in the liver that is responsible for iron transport in plasma. The structure of the molecule consists of two carbohydrate residues to which six sialic acid moieties can be attached. After periods of chronic, heavy alcohol consumption, carbohydrate-deficient transferrin (CDT) isoforms often increase, which makes CDT a useful marker in screening for alcohol abuse and monitoring progress of alcoholics in treatment. However, the precise mechanism behind CDT increase remains unknown. METHODS: A review of the most relevant literature on CDT was conducted with a computer-assisted literature search. RESULTS: During the past several years, a number of studies have explored possible mechanisms that may account for the alcohol-induced increase of CDT. An inhibition of protein synthesis and a general effect of alcohol on protein glycosylation have been reported. Although the exact mechanisms that underlie production of CDT are not yet fully understood, possible bases for the phenomenon are presented in this article. CONCLUSIONS: Experimental findings indicate that the ethanol-induced effect on glycoprotein metabolism is a multistep process in which protein transport and changes of enzyme activity may play an important role.

Alcoholism↗

The role of biomarkers in alcoholism medication trials.

BACKGROUND: Increasingly, biomarkers are being incorporated into the research design of clinical trials on medications to reduce drinking in alcoholics. To date, however, there has been little analysis of the unique roles that biomarkers can play in such investigations or of the practical and conceptual considerations that surround their best use in this context. METHODS: Clinical trials of alcoholism medications published between 1985 and the present were abstracted to determine how biomarkers were used and how changes in them related to self-report measures of drinking. RESULTS: Six uses of biomarkers were identified: determination of subjects to be included or excluded in the trial; description of baseline sample characteristics; primary and secondary outcome assessment; corroboration of self-reports of drinking status; specification of patients likely to respond to the medication; and evaluation of drug safety. CONCLUSION: Use of biomarkers in such studies appears warranted, particularly as an objective source of information on treatment efficacy that can be considered with patient self-report measures of drinking status. Biomarkers related to liver functioning also can assist in determination of drug safety for medications metabolized by the liver.

Alcoholism↗