Targeting Unique Epitopes on Highly Similar Proteins GDF-11 and GDF-8 with Modified DNA Aptamers

Area of Science:

• Biochemistry
• Molecular Biology
• Biotechnology

Background:

• Growth Differentiation Factor 11 (GDF-11) and GDF-8 are highly similar TGF-β family proteins.
• Distinguishing their specific biological functions is challenging due to cross-reactive binding reagents.

Purpose of the Study:

• To develop highly specific binding reagents for GDF-11 and GDF-8.
• To enable the elucidation of distinct biological roles for these closely related growth factors.

Main Methods:

• Utilized a combination of positive selection and counter-selection to identify specific aptamers.
• Employed Slow Off-rate Modified Aptamer (SOMAmer) technology for reagent development.
• Characterized binding affinity, specificity, and kinetics using various biochemical assays.

Main Results:

• Identified GDF-11 specific SOMAmer reagents with high affinity (0.05-1.2 nM K_d) and no cross-reactivity with GDF-8 (>1 μM K_d).
• Developed a GDF-8 specific SOMAmer with excellent affinity (0.23 nM K_d) and specificity.
• Demonstrated that standard positive selection alone yielded non-specific reagents, unlike the counter-selection method.
• Confirmed robust binding of aptamers across diverse assay conditions.

Conclusions:

• The developed SOMAmer reagents exhibit high affinity and specificity for GDF-11 and GDF-8.
• The selection method combining positive and counter-selection is crucial for differentiating highly similar proteins.
• These specific reagents are valuable tools for investigating the distinct biological functions of GDF-11 and GDF-8.