Conformation of dimeric apolipoprotein A-I milano on recombinant lipoprotein particles

Area of Science:

• Biochemistry
• Structural Biology
• Lipid Metabolism

Background:

• Apolipoprotein A-I (apoA-I) is crucial for high-density lipoprotein (HDL) structure and function.
• The naturally occurring apoA-I Milano (apoA-I(Milano)) mutation, characterized by a cysteine substitution, forms distinct homodimers.
• Wild-type apoA-I (apoA-I(WT)) forms defined relationships within phospholipid disks, influencing HDL conformation.

Purpose of the Study:

• To elucidate the conformational constraints of dimeric apoA-I(Milano) within recombinant HDL (rHDL) disks.
• To compare the structural organization of apoA-I(Milano) on rHDL with that of apoA-I(WT).
• To refine models of lipid-bound dimeric apoA-I(Milano) based on experimental distance constraints.

Main Methods:

• Utilized chemical cross-linking with lysine-selective homobifunctional cross-linkers on rHDL particles of 78 Å and 125 Å diameters.
• Employed mass spectrometry (MS/MS sequencing) to identify and confirm cross-linked peptides after reduction and trypsin digestion.
• Integrated cross-linking data to generate distance constraints for molecular modeling of lipid-bound apoA-I(Milano).

Main Results:

• A single apoA-I(Milano) dimer on 78 Å rHDL adopts a "belt" conformation, similar to apoA-I(WT), but with the C-terminus wrapping the particle periphery.
• Unlike apoA-I(WT), the C-terminal end of apoA-I(Milano) shields fatty acid chains from water.
• Two apoA-I(Milano) dimers on 125 Å rHDL do not encircle the disk but associate with a laminar micelle structure.

Conclusions:

• Dimeric apoA-I(Milano) exhibits distinct structural arrangements on rHDL compared to apoA-I(WT), influencing particle conformation.
• The unique C-terminal interaction of apoA-I(Milano) suggests altered lipid shielding mechanisms.
• These findings provide insights into the structural basis of apoA-I(Milano)'s function in lipid metabolism and cardiovascular health.