Abstract
The nerve growth factor (NGF) N-terminus peptide, NGF(1–14), and its acetylated form, Ac-NGF(1–14), were investigated to scrutinize the ability of this neurotrophin domain to mimic the whole protein. Theoretical calculations demonstrated that non-covalent forces assist the molecular recognition of TrkA receptor by both peptides. Combined parallel tempering/docking simulations discriminated the effect of the N-terminal acetylation on the recognition of NGF(1–14) by the domain 5 of TrkA (TrkA-D5). Experimental findings demonstrated that both NGF(1–14) and Ac-NGF(1–14) activate TrkA signaling pathways essential for neuronal survival. The NGF-induced TrkA internalization was slightly inhibited in the presence of Cu2+ and Zn2+ ions, whereas the metal ions elicited the NGF(1–14)-induced internalization of TrkA and no significant differences were found in the weak Ac-NGF(1–14)-induced receptor internalization. The crucial role of the metals was confirmed by experiments with the metal-chelator bathocuproine disulfonic acid, which showed different inhibitory effects in the signaling cascade, due to different metal affinity of NGF, NGF(1–14) and Ac-NGF(1–14). The NGF signaling cascade, activated by the two peptides, induced CREB phosphorylation, but the copper addition further stimulated the Akt, ERK and CREB phosphorylation in the presence of NGF and NGF(1–14) only. A dynamic and quick influx of both peptides into PC12 cells was tracked by live cell imaging with confocal microscopy. A significant role of copper ions was found in the modulation of peptide sub-cellular localization, especially at the nuclear level. Furthermore, a strong copper ionophoric ability of NGF(1–14) was measured. The Ac-NGF(1–14) peptide, which binds copper ions with a lower stability constant than NGF(1–14), exhibited a lower nuclear localization with respect to the total cellular uptake. These findings were correlated to the metal-induced increase of CREB and BDNF expression caused by NGF(1–14) stimulation. In summary, we here validated NGF(1–14) and Ac-NGF(1–14) as first examples of monomer and linear peptides able to activate the NGF-TrkA signaling cascade. Metal ions modulated the activity of both NGF protein and the NGF-mimicking peptides. Such findings demonstrated that NGF(1–14) sequence can reproduce the signal transduction of whole protein, therefore representing a very promising drug candidate for further pre-clinical studies.
Highlights
Neurotrophins are a family of structurally conserved growth factors involved in differentiation, survival of neurons (Huang and Reichardt, 2001; Chao, 2003) as well as of non-neuronal cell type (Sofroniew et al, 2001; Reichardt, 2006; Caporali and Emanueli, 2009)
With the simulations carried out for the wildtype nerve growth factor (NGF)(1–14), parallel tempering simulations were run in parallel, in order to sample the conformations of Ac-NGF(1– 14) at physiological pH
We recently reported that NGF(1–14), a linear peptide encompassing the 1–14 sequence of the human NGF protein, is able to activate the TrkA pathway, to induce the phosphorylation of cAMP response element-binding protein (CREB) Ser-133, and to affect the PC12 proliferation rate (Travaglia et al, 2015)
Summary
Neurotrophins are a family of structurally conserved growth factors involved in differentiation, survival of neurons (Huang and Reichardt, 2001; Chao, 2003) as well as of non-neuronal cell type (Sofroniew et al, 2001; Reichardt, 2006; Caporali and Emanueli, 2009). They include nerve growth factor (NGF), brain derived neurotrophic factor (BDNF), NT-3 (neurotrophin 3), and NT-4. NGF, composed of 118 amino acid residues (Angeletti et al, 1971) has been discovered in the 1950s (Levi-Montalcini and Hamburger, 1951; Levi-Montalcini, 1952; Cohen et al, 1954; Levi-Montalcini and Booker, 1960); it displays multiple physiological actions in the central nervous system, showing neurotrophic effects and resulting critical for the neurite outgrowth and survival and maintenance of neurons (Hu et al, 2005; Tucker et al, 2008; Xu et al, 2012).
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
