Intrinsic structural disorder of mouse proNGF

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 p75NTR receptors. SAXS measurements revealed the rm-proNGF to be dimeric and anisometric, with the pro-peptide domain being intrinsically unstructured. Ab initio reconstructions assuming two-fold 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 co-existence of different conformations contributing to the experimental scattering pattern, with no symmetry constraints, suggests the crab-like to be a more likely proNGF conformation. In order 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 pro-peptide of proNGF may act as an intrinsically unstructured protein domain, characterized by a molecular promiscuity in the interaction/binding to multiple partners (TrkA and p75NTR 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.