last update: March 21, 2001
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STC-ERSP Program Details Principal Investigator: R.M. Kelly Project Title: Enzymes from extremophiles in CO2 (Former Prog #24) Research Plan Connectivity Outreach Components Requested Budget Allocation - Year 1 Plans for Additional Funding Research Plan Overall objectives This project focuses on the stability and activity of hyperthermophilic enzymes in nonaqueous and partially aqueous media, with particular focus on CO2. Particular focus is on hyperthermophilic esterases and lipases as catalysts for a variety of synthetic biotransformations. We intend to answer several questions concerning the stability and activity of these enzymes in liquid and supercritical CO2, including the influence of pressure and temperature on enzyme kinetics and product formation, the impact of co-solvents on enzyme function, and the strategic opportunities offered by the use of stable and active biocatalysts in CO2 in comparison to organic solvents and water. Relation to overall objectives of the Center Although there are many advantages offered by the use of organic solvents as reaction media for biotransformations (unusual selectivities, altered solubility of reactants and products), use of an environmentally benign, nonaqueous solvent would be even more attractive. Liquid and supercritical CO2 offers such an alternative. There has been relatively little work done concerning biocatalysis in CO2 and none involving enzymes from extremophilic microorganisms. CO2 offers the additional advantage of significantly simplifying the product recovery step through depressurization. Approach and Year 1-Year 5 timelines Years 1-2: Perform initial studies using a thermally stable esterase from the hyperthermophilic archaeon Sulfolobus solfataricus. Enzyme kinetics will be done in several reaction mileau: water, water-cosolvent mixtures, organic solvents, liquid CO2 and supercritical CO2. Several esterification reactions will be considered in addition to the stability characteristics of the enzyme. Techniques for immobilizing the enzyme will also be studied. Years 2-5: Several other hyperthermophilic esterases and lipases will be examined for their ability to catalyze reactions in organic solvents and CO2. The emphasis here will be reactions of industrial significance including the synthesis and separation of pharmaceutical intermediates. These enzymes will be obtained through existing collaborations as well as by PCR-cloning from genomic data corresponding to completed hyperthermophile genome sequences. Thrust area of this proposal Thrust Area D: Chemistry and Catalysis Connectivity Collaborators, multi-institutional, multi-disciplinary components We will collaborate with Ruben Carbonell and Joe DeSimone on the chemistry of the reactions to be studied, techniques for immobilizing enzymes using polymers that can be made to swell under controlled conditions, and on the use of cosolvents and surfactants in reaction media formulations. Related work in other thrust areas The potential scale-up of these enzyme reaction systems could involved collaborations with those working the Rate Processes thrust area. Sharing of resources (students, supplies, equipment, etc.) We will need access to certain equipment not present in our laboratory. This includes a CO2reaction cell, GC/MS, and other analytical equipment. The postdoc assigned to the project, Hong Chen, will likely spend time in both our lab and Joe DeSimone's lab. The same is true to a lesser extent for the graduate student, David Sehgal. Outreach Components Suggested K-12 Outreach Ideas Kelly would be available to speak about extremophilic organisms and enzymes to high school science classes. The possibility of hosting a high school student in our lab also exists. Requested Budget Allocation - Year 1 Personnel salaries
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