Nerve growth factor: Structure/function relationships

Abstract

Nerve growth factor (NGF), which has a tertiary structure based on a cluster of 3 cystine disulfides and 2 very extended, but distorted 0-hairpins, is the prototype of a larger family of neurotrophins. Prior to the availability of cloning techniques, the mouse submandibular gland was the richest source of NGF and provided sufficient material to enable its biochemical characterization. It binds as a dimer to at least 2 cell-surface receptor types expressed in a variety of neuronal and non-neuronal cells. Residues involved in these interactions and in the maintenance of tertiary and quaternary structure have been identified by chemical modification and site-directed mutagenesis, and this information can be related to their location in the 3-dimensional structure. For example, interactions between aromatic residues contribute to the stability of the NGF dimer, and specific surface lysine residues participate in receptor contacts. The conclusion from these studies is that receptor interactions involve broad surface regions, which may be composed of residues from both protomers in the dimer.Keywords: nerve growth factor; neurotrophins; structure/function relationships The elucidation of the structure/function relationships of a protein requires a precise knowledge of the 3-dimensional structure and the identification of the residues that participate in biological activity. The latter were traditionally defined by chemical modification, which has now been supplanted, to a large degree, by site-directed mutagenesis, primarily because it offers the possibility of making substitutions anywhere in the molecule. However, despite its limitations, chemical modification does offer 1 major advantage: it is carried out on the intact protein and does not generally require refolding, a problem that has hampered the preparation of many recombinant derivatives. As is often the case, both approaches used in concert may provide much more information than either alone.

A New Nerve Growth Factor-Mimetic Peptide Active on Neuropathic Pain in Rats

“…Many efforts have been devoted to design mimetics using NGF fragments. Crystallographic and molecular modeling studies of NGF/TrkA complexes (McDonald et al, 1991;McDonald and Chao, 1995;Urfer et al, 1998;Wiesmann et al, 1999), mutagenesis analysis (Ibànez et al, 1992;Kahle et al, 1992;Drinkwater et al, 1993;Shih et al, 1994;Woo et al, 1995;Krüttgen et al, 1997;Kullander et al, 1997;Rydén and Ibàñez, 1997), and chemical modification of specific amino acids (Bradshaw et al, 1994;McDonald and Chao, 1995) have indicated the hydrophilic loops 1 and 4 and the N-terminal region of NGF as the most relevant for its biological activity. Among various peptides reported in the literature, linear peptides showed inhibitory, but no agonist activity (Longo et al, 1990), and cyclic peptides corresponding to the loop 1 sequence were shown later to have a 10-fold higher inhibitory activity (LeSauteur et al, 1995).…”

Nerve Growth Factor (NGF) Loop 4 Dimeric Mimetics Activate ERK and AKT and Promote NGF-like Neurotrophic Effects

“…Recombinant substitution studies indicated that residues Lys 32 and Lys 34 (17) and other residues (18) are likely to interact with p75 receptors. The NGF sites interacting with TrkA have also been derived via chemical modification (11,19,20), recombinant protein (18, 20 -23), and NGF-TrkA co-crystallization approaches (24). Taken together, these studies indicate that TrkA binding sites consist of residues in NGF loop 2 (residues 40 -49), loop 4 (residues 91-97), the N terminus (residues [1][2][3][4][5][6][7][8], and the C terminus (residues 111-115).…”

“…Multiple techniques have been used to deduce which domains of the NGF protein interact with NGF receptors (11). A peptide mapping approach in which synthetic peptides with sequences corresponding to specific NGF regions were tested for their ability to inhibit NGF activity pointed to residues 29 -35 as a key active site (12).…”

Two Restricted Sites on the Surface of the Nerve Growth Factor Molecule Independently Determine Specific TrkA Receptor Binding and Activation

“…A first approach used chemical modification of NGF where residues receptive for different agents could be blocked and the effect analyzed (for review, see Ref. 19). Enzymatic cleavage of NGF has identified the NH 2 terminus to be critical for binding to TrkA (20 -22).…”