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Ithaca, NY, United States

Bisharyan Y.,Cornell University | Bisharyan Y.,Tetragenetics | Clark T.G.,Cornell University
Mitochondrion | Year: 2011

Here we demonstrate that ciliated protozoa can jettison mitochondria as intact organelles, releasing their contents to the extracellular space either in a soluble form, or in association with membrane vesicles at the cell periphery. The response is triggered by lateral clustering of GPI-anchored surface antigens, or by heat shock. In the first instance, extrusion is accompanied by elevated levels of intracellular calcium and is inhibited by Verapamil and BAPTA-AM arguing strongly for the involvement of calcium in triggering the response. Cells survive mitochondrial discharge raising the interesting possibility that extrusion is an early evolutionary adaptation to cell stress. © 2011 Elsevier B.V. and Mitochondria Research Society.

This invention is directed to methods for the production of immunogenic granular particles. In certain embodiments, the invention is directed to methods and products for the production immunogenic granular particles produced in ciliates. In certain embodiments, the invention is directed to compositions comprising Grl/Ag fusion polypeptides.

Agency: Department of Defense | Branch: Army | Program: STTR | Phase: Phase II | Award Amount: 750.00K | Year: 2007

Recombinant proteins are critically important in a wide range of applications that extend from the treatment of human and animal disease, to chemical and biological defense. Production of genetically engineered vaccine antigens, therapeutic proteins (including monoclonal antibodies), industrial enzymes, biopolymers, and bioremediation agents now constitutes a multibillion dollar-per-year industry. However, the need for low cost systems for the rapid production of these proteins remains great. In Phase I we demonstrated feasibility of the use of a free-living ciliate, Tetrahymena thermophila, as novel platform for high-level expression of the H5 hemagglutinin of avian influenza virus. In Phase II we will extend this work to commercially relevant enzymes, and antibody fragments targeted against biowarfare agents with the goal of benchmarking both yield and production costs against competing systems. In addition, we will develop user-friendly protein expression kits for the research tool market based on the TetraExpressTM platform.

Agency: Department of Defense | Branch: Army | Program: STTR | Phase: Phase I | Award Amount: 100.00K | Year: 2006

To fully realize the benefits of genetically engineered proteins, manufacturing costs and development time must be significantly reduced. Technologies that combine robust expression, and inexpensive methods of purification are key to this goal. This Phase I proposal will demonstrate the feasibility of Tetrahymena thermophila, a common pondwater ciliate, as a novel system for low cost production of recombinant proteins using strategies that focus on a “model” vaccine antigen with considerable importance in human and animal health, namely, the hemagglutinin protein (HA) of influenza virus. Our specific goals are to 1) introduce the HA gene from the current H5N1 strain of avian flu into T. thermophilia 2) screen recombinant cell lines for the expression of HA using antibodies specific for the protein; and 3) devise rapid, inexpensive methods for downstream purification of candidate vaccine antigens.

Agency: Department of Health and Human Services | Branch: | Program: STTR | Phase: Phase I | Award Amount: 99.13K | Year: 2006

DESCRIPTION (provided by applicant): These studies are intended to establish Tetrahymena as an alternative expression system for use in high-throughput structural studies, and for large-scale production of membrane proteins with importance in human health. Towards that end, we will attempt to express 3 model membrane proteins in this system using a unique and versatile high-copy number cloning vector. The vector (now under construction) takes advantage of a novel biological feature of Tetrahymena in which the ribosomal genes are amplified to ~ 18,000 copy number as part of an autonomously replicating palindromic minichromosome. Using a similar vector that lacks a multicloning site and affinity tags, we have been able to express a heterologous GPI-anchored membrane protein in Tetrahymena to levels greater than 3% of total cell protein. We now want to validate this approach for important transmembrane proteins in humans. This includes the human epidermal growth factor receptor (a G-protein coupled receptor) and the ionotropic glutamate receptor GluRG (which acts as a channel protein). Tetrahymena has a number of distinct features which set it apart from more conventional protein expression systems and which make it ideal as a vehicle for the production of recombinant eucaryotic membrane proteins. A successful outcome, in this case overexpression of functional proteins, will provide proof-of-principal for the use of this system and make it an attractive tool for use by the biopharmaceutical industry. Our Small Business Partner, Tetragenetics Inc, seeks to commercialize on this technology and make it available to researchers in both the academic and industry arenas through protein expression kits and contract expression services. Eucaryotic membrane proteins contribute to an array of vital cell functions and are essential to human health. The technology described here is intended to better understand how these proteins function, leading ultimately to strategies for the treatment of serious disorders ranging from cancer to stoke.

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