title: Intrinsic structural disorder of mouse proNGF.
abstract: The unprocessed precursor of the Nerve Growth Factor (NGF), proNGF, has additional functions, besides its initially described role as a chaperone for NGF folding. The precursor protein endows apoptotic and/or neurotrophic properties, in contrast to the mature part. The structural and molecular basis for such distinct activities are presently unknown. Aiming to gain insights into the specific molecular interactions that govern rm-proNGF biological activities versus those of its mature counterpart, a structural study by synchrotron small angle X-ray scattering (SAXS) in solution was carried out. The different binding properties of the two proteins were investigated by surface plasmon resonance (SPR) using, as structural probes, a panel of anti-NGF antibodies and the soluble forms of TrkA and p75(NTR) receptors. SAXS measurements revealed the rm-proNGF to be dimeric and anisometric, with the propeptide domain being intrinsically unstructured. Ab initio reconstructions assuming twofold symmetry generated two types of structural models, a globular "crab-like" and an elongated shape that resulted in equally good fits of the scattering data. A novel method accounting for possible coexistence of different conformations contributing to the experimental scattering pattern, with no symmetry constraints, suggests the "crab-like" to be a more likely proNGF conformation. To exploit the potential of chemical stabilizers affecting the existing conformational protein populations, SAXS data were also collected in the presence of ammonium sulphate. An increase of the proNGF compactness was observed. SPR data pinpoints that the propeptide of proNGF may act as an intrinsically unstructured protein domain, characterized by a molecular promiscuity in the interaction/binding to multiple partners (TrkA and p75(NTR) receptors and a panel of neutralizing anti-NGF antibodies) depending on the physiological conditions of the cell. These data provide a first insight into the structural basis for the selectivity of mouse short proNGF, versus NGF, towards its binding partners.
authors.affiliation: SISSA-ISAS, Building Q1, Area Science Park - Basovizza, S.S 14 Km 163.5, 34012 Trieste, Italy. f.paoletti@ebri.i
authors.forename: Francesca
authors.initials: F
authors.lastname: Paoletti
authors.forename: Sonia
authors.initials: S
authors.lastname: Covaceuszach
authors.forename: Peter V
authors.initials: PV
authors.lastname: Konarev
authors.forename: Stefania
authors.initials: S
authors.lastname: Gonfloni
authors.forename: Francesca
authors.initials: F
authors.lastname: Malerba
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Schwarz
authors.forename: Dmitri I
authors.initials: DI
authors.lastname: Svergun
authors.forename: Antonino
authors.initials: A
authors.lastname: Cattaneo
authors.forename: Doriano
authors.initials: D
authors.lastname: Lamba
mesh_terms: D000595:Amino Acid Sequence; D000645:Ammonium Sulfate / Q000737:chemistry; D000818:Animals; D000915:Antibody Affinity; D003198:Computer Simulation; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D020932:Nerve Growth Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016716:PC12 Cells; D010766:Phosphorylation; D011487:Protein Conformation; D011498:Protein Precursors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D020917:Receptor, trkA / Q000737:chemistry / Q000378:metabolism*; D017475:Receptors, Nerve Growth Factor / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D053838:Scattering, Small Angle; D016415:Sequence Alignment; D020349:Surface Plasmon Resonance; D014961:X-Ray Diffraction
chemical_list: D011498:Protein Precursors; D017475:Receptors, Nerve Growth Factor; D011994:Recombinant Proteins; C517634:Ngfr protein, mouse; C489226:pro-nerve growth factor, mouse; D020932:Nerve Growth Factor; D020917:Receptor, trkA; D000645:Ammonium Sulfate