Characterization and structural analysis of a functional human serum transferrin variant and implications for...

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Human serum transferrin (hTf) is crucial for iron transport.
• A single known functional variant of hTf exhibits altered properties.
• Understanding this variant's molecular basis is key to its function.

Purpose of the Study:

• To characterize the molecular basis of the functional human serum transferrin variant.
• To investigate the structural and metal-binding consequences of the identified mutation.
• To elucidate the mechanism behind the variant's reduced receptor affinity.

Main Methods:

• Peptide sequencing after cyanogen bromide digestion.
• Genomic PCR coupled with cycle sequencing.
• UV difference spectroscopy, protein fluorescence quenching, and solution X-ray scattering.

Main Results:

• Identified a Glycine to Arginine substitution at position 394 (Gly394Arg) due to a G to A nucleotide transition.
• The variant binds Zn(2+), Al(3+), and Cu(2+) to only one of its two metal-binding sites.
• Structural analysis revealed the C-lobe remains open in the iron-bound state due to a salt bridge involving Arg394, explaining reduced receptor affinity.

Conclusions:

• The Gly394Arg substitution is the molecular cause of the functional hTf variant.
• The mutation disrupts normal metal-binding site stoichiometry and alters protein conformation.
• The 'open' C-lobe conformation likely underlies the reduced affinity for the transferrin receptor.