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EPISODEs goal is to maximize the regional benefits of Structural Biology Research Infrastructures in NMR and associated technologies for the economic development of the pharma/biotech industries in Tuscany, Berlin-Brandenburg and beyond. As traditional Structural Biology research moves towards new horizons, a major goal has become a systemic view of Life, implying a change of focus from single molecules and interactions to an integrated view of networks of interactions at varying levels of biological organization. NMR and associated technologies are an integral part of such research, and their importance has recently been reflected in the ESFRI Roadmaps INSTRUCT Integrated Structural Biology Infrastructure. While this emerging paradigm presents myriad new opportunities for the R&D programs of the pharma/biotech sectors, the technologies and their associated research infrastructures have not yet been fully exploited, a situation that must urgently be understood and rectified. Berlin-Brandenburg and Tuscany are uniquely poised to meet this challenge, in part through the presence of strong research infrastructures. EPISODE will encourage cooperation and collaboration among regional research institutions, business and areas agencies to couple research with revenue, through the following actions: (i) research-driven clusters will be expanded and stimulated interregionally and transnationally; (ii) current regional capabilities in terms of research potential and exploitation will be studied; (iii) a Joint Action Plan will be developed to define how to drive future economic development. Dissemination and outreach efforts, the encouragement of collaborations, support measures for SMEs and spin-offs, and the mentoring of less-developed regions will be given a special place throughout the project. EPISODE will positively impact research potential and economic potential, with corresponding effects on the health and welfare of regional, European and global populations.


Merk H.,RiNA Netzwerk RNA Technologien GmbH | Rues R.-B.,Goethe University Frankfurt | Gless C.,RiNA Netzwerk RNA Technologien GmbH | Beyer K.,RiNA Netzwerk RNA Technologien GmbH | And 4 more authors.
Biological Chemistry | Year: 2015

G protein-coupled receptors, like many other membrane proteins, are notoriously difficult to synthesize in conventional cellular systems. Although expression in insect cells is considered the preferred technique for structural characterizations in particular, inefficient membrane translocation, instability, toxic effects and low yields still pose clear limitations for their production in living cells. Recent studies started to explore alternative strategies for the in vitro production of problematic membrane proteins in cell-free lysates in combination with supplied membranes. We provide a detailed study on the production efficiencies and quality of G protein-coupled receptors, Fab fragments and other proteins synthesized in insect cell lysates containing endogenous microsomes. Effects of different reaction kinetics, redox conditions and sample preparations on the specific activities of synthesized proteins have been analyzed. The extent of glycosylation, membrane translocation and percentages of ligand binding active fractions of synthesized protein samples have been determined. We provide strong evidence that membrane insertion of integral membrane proteins can represent a prime limiting factor for their preparative scale in vitro production. Improved expression protocols resulting into higher production rates yielded more active protein in case of Fab fragments, but not in case of the human endothelin B receptor. © 2015 by De Gruyter.


Patent
Rina Netzwerk Rna Technologien Gmbh | Date: 2014-03-24

The invention relates to a method for producing a lysate used for cell-fee protein biosynthesis, comprising the following steps: a) a genomic sequence in an organism, which codes for an essential translation product that reduces the yield of cell-fee protein biosynthesis, is replaced by the foreign DNA located under a suitable regulatory element, said foreign DNA coding for the essential translation product that additionally contains a marker sequence; b) the organism cloned according to step a) is cultivated; c) the organisms from the culture obtained in step b) are lysed; and d) the essential translation product is eliminated by means of a separation process that is selective for the marker sequence. Also discussed are said lysate and the use thereof


Patent
Rina Netzwerk Rna Technologien Gmbh | Date: 2012-04-10

The invention relates to a method for preparative in vitro protein synthesis in a cell-free transcription/translation system, comprising the following steps: a) in a reaction vessel, a reaction solution is prepared, comprising the following synthesis substances: components of the transcription/translation apparatus for a defined-protein, amino acids, and metabolic components supplying energy and being necessary for the synthesis of the defined protein, b) the synthesis is performed in the reaction vessel in a defined period of time, c) after expiration of the defined period of time, the reaction solution is subjected to a separation step, in which generated low-molecular metabolic products are separated from the solution (and extracted).


Patent
Rina Netzwerk RNA Technologien GmbH | Date: 2014-01-10

The invention relates to a method for preparative in vitro protein synthesis in a cell-free transcription/translation system, comprising the following steps: a) in a reaction vessel, a reaction solution is prepared, comprising the following synthesis substances: components of the transcription/translation apparatus for a defined-protein, amino acids, and metabolic components supplying energy and being necessary for the synthesis of the defined protein, b) the synthesis is performed in the reaction vessel in a defined period of time, c) after expiration of the defined period of time, the reaction solution is subjected to a separation step, in which generated low-molecular metabolic products are separated from the solution (and extracted).


Patent
PLS Design GmbH and RiNA NETZWERK RNA TECHNOLOGIEN GMBH | Date: 2013-11-07

The invention relates to a method for the sequence information of PNA molecules of a specific PNA molecule species, wherein, the PNA molecules are contacted with different nucleic acid molecule species comprising nucleic acid molecules with nucleotides, wherein the nucleic acid molecules partially comprise a nucleic acid sequence that is complementary to a partial sequence of the PNA molecule, wherein nucleic acid molecules having complementary sequences bind to the PNA molecules forming nucleic acid/PNA hybrids, wherein nucleic acid molecules with non-complementary sequences are separated from the hybrids, wherein thereafter the hybrids are dissociated into single stranded hybrid nucleic acid molecules and PNA molecules, wherein the single stranded hybrid nucleic acid molecules are subjected to a sequencing process providing hybrid sequence information about the single stranded hybrid nucleic acid sequence, and wherein the hybrid sequence information is optionally translated into the complementary PNA sequence information.


Patent
Rina Netzwerk Rna Technologien Gmbh | Date: 2010-01-08

The invention relates to a method for preparative in vitro protein synthesis of an expression product in a cell-free transcription/translation system, comprising the following steps: a) in a reaction vessel, a reaction solution is prepared, comprising the following synthesis substances: components of the transcription/translation apparatus for a defined expression product, amino acids, and metabolic components supplying energy and being necessary for the synthesis of the defined protein, b) the synthesis is performed in the reaction vessel in a defined period of time, without separating generated substances and without adding consumed synthesis substances within the defined period of time, c) after expiration of the defined period of time, the reaction solution is subjected to a separation step, in which generated low-molecular metabolic products and/or reaction inhibitors are separated from the solution (and extracted), d) immediately before, after or at the same time as step c) consumed synthesis substances are supplemented, e) steps b), c) and d) are repeated at least once with the reaction solution of step d), and at the last execution of step b) steps c) and d) may be left out.


Patent
Rina Netzwerk Rna Technologien Gmbh | Date: 2013-04-19

The invention relates to a method for preparative in vitro protein synthesis in a cell-free transcription/translation system, comprising the following steps: a) in a reaction vessel, a reaction solution is prepared, comprising the following synthesis substances: components of the transcription/translation apparatus for a defined-protein, amino acids, and metabolic components supplying energy and being necessary for the synthesis of the defined protein, b) the synthesis is performed in the reaction vessel in a defined period of time, c) after expiration of the defined period of time, the reaction solution is subjected to a separation step, in which generated low-molecular metabolic products are separated from the solution (and extracted).


Patent
Rina Netzwerk Rna Technologien Gmbh | Date: 2012-09-28

The invention relates to a method for producing a lysate used for cell-fee protein biosynthesis, comprising the following steps: a) a genomic sequence in an organism, which codes for an essential translation product that reduces the yield of cell-fee protein biosynthesis, is replaced by the foreign DNA located under a suitable regulatory element, said foreign DNA coding for the essential translation product that additionally contains a marker sequence; b) the organism cloned according to step a) is cultivated; c) the organisms from the culture obtained in step b) are lysed; and d) the essential translation product is eliminated by means of a separation process that is selective for the marker sequence. Also discussed are said lysate and the use thereof.


Merk H.,RiNA Netzwerk RNA Technologien GmbH | Gless C.,RiNA Netzwerk RNA Technologien GmbH | Maertens B.,Qiagen | Gerrits M.,RiNA Netzwerk RNA Technologien GmbH | Stiege W.,RiNA Netzwerk RNA Technologien GmbH
BioTechniques | Year: 2012

A eukaryotic cell-free system based on Spodoptera frugiperda cells was developed for the convenient synthesis of Fab antibody fragments and other disulfide bridge containing proteins. The system uses (i) a cell lysate that is mildly prepared under slightly reduced conditions, thus maintaining the activity of vesicles derived from the endoplasmic reticulum, (ii) signal peptide dependent translocation into these vesicles, and (iii) a redox potential based on reduced and oxidized glutathione. Monomeric heavy and light immunoglobulin chains are almost completely converted to highly active dimeric Fab joined by intermolecular disulfide bridges without supplementation of chaperones or protein disulfide isomerase. The applicability of the system is demonstrated by the synthesis of anti-lysozyme and anti-CD4 Fab antibody fragments yielding approximately 10 μg Fab per milliliter reaction mixture. The lack of endotoxins in this system is a prerequisite that synthesized Fab can be applied directly using whole synthesis reactions in cell-based assays that are sensitive to this substance class. Moreover, the system is compatible with PCR-generated linear templates enabling automated generation of antibody fragments in a high-throughput manner, and facilitating its application for screening and validation purposes.

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