Linker Length and Composition within Disordered Binding Motifs Modulates the Avidity and Reversibility of a...

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• Intrinsically disordered proteins regulate the cellular transcriptional response to hypoxia.
• Hypoxia-inducible factor (HIF)-1α is a key regulator of oxygen stress genes.
• CITED2 acts as a feedback inhibitor, terminating the hypoxic response.

Purpose of the Study:

• To investigate the role of binding cooperativity in the unidirectional hypoxic switch.
• To understand how linker mutations in HIF-1α affect its binding to TAZ1 and competition with CITED2.
• To elucidate the molecular basis for tuning the hypoxic switch.

Main Methods:

• Mutagenesis of the linker sequence in the HIF-1α activation domain.
• Analysis of thermodynamic coupling between TAZ1-binding motifs.
• Assessment of HIF-1α binding affinity and competition with CITED2.

Main Results:

• Linker mutations enhancing HIF-1α affinity improved its competition with CITED2.
• Binding affinity is influenced by linker flexibility (glycine residues) and electrostatic interactions (arginine side chain).
• Mutants provided insights into the molecular basis of the hypoxic switch's tuning.

Conclusions:

• Linker sequence and flexibility are critical for regulating HIF-1α binding affinity.
• Sub-optimal HIF-1α binding affinity, driven by evolutionary pressure, ensures the unidirectional hypoxic switch.
• Understanding these mechanisms is key to regulating oxygen stress responses.