Comparative equilibrium denaturation studies of the neurotrophins: nerve growth factor, brain-derived neurotrophic factor, neurotrophin 3, and neurotrophin 4/5.


The neurotrophins are a family of small dimeric proteins required for the development and survival of vertebrate neurons. Solvent denaturation studies were used to compare recombinant human nerve growth factor (hNGF), brain-derived neurotrophic factor (BDNF), neurotrophin 3 (NT-3), and neurotrophin 4/5 (NT-4/5) to nerve growth factor isolated from mouse submaxillary glands (mNGF). Although greater than 50% sequence identity is conserved among this family, significant structural differences were revealed by the folding and unfolding of these proteins. Denaturation in guanidine hydrochloride and renaturation at pH 7 and 3.5 were monitored by fluorescence intensity, fluorescence polarization, and circular dichroism. The midpoint of equilibrium unfolding curves for all four neurotrophins was independent of the technique but was dependent on protein concentration, indicating that a two-state model involving native neurotrophin dimers and denatured neurotrophin monomers (N2 = 2D) describes the equilibrium between folded and unfolded neurotrophins. The conformational stabilities of the dimeric neurotrophins revealed that mNGF had the lowest conformational stability (19.3 kcal/mol); hNGF, NT-3, and NT-4/5 had intermediate stabilities, and BDNF had the highest stability (26.4 kcal/mol). Recovery of native spectroscopic characteristics upon removal of denaturant indicated that the unfolding process is reversible. Accordingly, unfolding and refolding curves were coincident for mNGF or NT-4/5 at pH 7 and 3.5 and for BDNF at pH 3.5. However, BDNF and NT-3 unfolding and refolding curves were not coincident at pH 7. The stability of the neurotrophins decreased as pH decreased, with compact monomeric intermediates (N2 = [2I] = 2D) becoming populated below pH 4. The differences in stability, pH dependence, and coincidence of refolding curves distinguish the homologous structures of the neurotrophins. Study holds ProTherm entries: 4432, 4433, 4434, 4435, 4436, 4437, 4438, 4439 Extra Details: dimeric proteins; two-state model; conformational stability;,refolding curves

Submission Details

ID: wZu6xm744

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:25 p.m.

Version: 1

Publication Details
Timm DE;de Haseth PL;Neet KE,Biochemistry (1994) Comparative equilibrium denaturation studies of the neurotrophins: nerve growth factor, brain-derived neurotrophic factor, neurotrophin 3, and neurotrophin 4/5. PMID:8161524
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)

Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
1B8K 1999-02-01T00:00:00+0000 2.15 Neurotrophin-3 from Human
1NT3 1999-05-17T00:00:00+0000 2.4 HUMAN NEUROTROPHIN-3
3BUK 2008-01-02T00:00:00+0000 2.6 Crystal Structure of the Neurotrophin-3 and p75NTR Symmetrical Complex

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Neurotrophin-3 P20181 NTF3_MOUSE
99.2 Neurotrophin-3 P18280 NTF3_RAT
95.7 Neurotrophin-3 P20783 NTF3_HUMAN
94.6 Neurotrophin-3 Q9TST2 NTF3_FELCA
91.5 Neurotrophin-3 Q06AV0 NTF3_PIG
92.7 Neurotrophin-3 Q08DT3 NTF3_BOVIN