http://www.cnr.it/ontology/cnr/individuo/prodotto/ID188855

Direct intracellular selection and biochemical characterization of a recombinant anti-proNGF single chain antibody fragment. (Articolo in rivista)

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Direct intracellular selection and biochemical characterization of a recombinant anti-proNGF single chain antibody fragment. (Articolo in rivista) (literal)

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Paoletti F., Malerba F., Konarev P.V., Visintin M., Scardigli R., Fasulo L., Lamba D., Svergun D.I., Cattaneo A. (2012)
Direct intracellular selection and biochemical characterization of a recombinant anti-proNGF single chain antibody fragment.
in Archives of biochemistry and biophysics (Online); Elsevier Inc., San Diego (Stati Uniti d'America)
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Paoletti F., Malerba F., Konarev P.V., Visintin M., Scardigli R., Fasulo L., Lamba D., Svergun D.I., Cattaneo A. (literal)

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Paoletti F. - EBRI - European Brain Research Institute, Via del Fosso di Fiorano 64, 00143 Rome, Italy; Malerba F. - EBRI - European Brain Research Institute, Via del Fosso di Fiorano 64, 00143 Rome, Italy; and Scuola Normale Superiore, Piazza dei Cavalieri 7, 56126 Pisa, Italy; Konarev P.V. - European Molecular Biology Laboratory, Hamburg Outstation, Notkestrasse 85, 22603 Hamburg, Germany; Visintin M. - Rottapharm Biotech s.r.l., Area Science Park - Basovizza, S.S. 14, Km 163.5, 34149 Trieste, Italy; Scardigli R. - Istituto di Farmacologia Traslazionale, CNR, Via del Fosso del Cavaliere 100, 00133 Roma, Italy; and EBRI - European Brain Research Institute, Via del Fosso di Fiorano 64, 00143 Rome, Italy; Fasulo L. - EBRI - European Brain Research Institute, Via del Fosso di Fiorano 64, 00143 Rome, Italy; Lamba D. - Istituto di Cristallografia, CNR, Area Science Park - Basovizza, S.S. 14, Km 163.5, 34149 Trieste, Italy; Svergun D.I. - European Molecular Biology Laboratory, Hamburg Outstation, Notkestrasse 85, 22603 Hamburg, Germany; Cattaneo A. - EBRI - European Brain Research Institute, Via del Fosso di Fiorano 64, 00143 Rome, Italy; and Scuola Normale Superiore, Piazza dei Cavalieri 7, 56126 Pisa, Italy (literal)

Titolo

Direct intracellular selection and biochemical characterization of a recombinant anti-proNGF single chain antibody fragment. (literal)

Abstract

proNGF, the precursor of the neurotrophin NGF, is widely expressed in central and peripheral nervous system. Its physiological functions are still largely unknown, although it emerged from studies in the last decade that proNGF has additional and distinct functions with respect to NGF, besides acting chaperonelike for NGF folding during its biogenesis. The regulation of proNGF/NGF ratio represents a crucial process for homeostasis of brain and other tissues, and understanding the molecular aspects of these differences is important. We report the selection and characterization of a recombinant monoclonal anti-proNGF antibody in single chain Fv fragment (scFv) format. The selection exploited the Intracellular Antibody Capture Technology (IACT), starting from a naïve mouse SPLINT (Single Pot Library of INTracellular antibodies) library. This antibody (scFv FPro10) was expressed recombinantly in Escherichia coli, was proven to be highly soluble and stable, and thoroughly characterized from the biochemical-biophysical point of view. scFv FPro10 displays high affinity and specificity for proNGF, showing no cross-reactivity with other pro-neurotrophins. A structural model was obtained by SAXS. scFv FPro10 represents a new tool to be exploited for the selective immunoanalysis of proNGF, both in vitro and in vivo, and might help in understanding the molecular function of proNGF in neurodegeneration. (literal)

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