Actin-related protein (Arp2) inserts into artificial lipid membranes

Arp2 is localized in the cytoplasm of eukaryotic cells where it controls actin dynamics. Computer analyses have suggested one possible lipid binding site, residues 185-202 of the primary amino acid sequence on Arp2, that could allow for membrane attachment/insertion. We expressed this region as a fusion protein with schistosomal glutathione S-transferase (GST) and investigated the interaction of this fragment with mixtures of dioleoylphosphatidylserine (DOPS) and dioleoylphosphatidylglycerol (DOPG) phospholipids in reconstituted lipid bilayers using differential scanning calorimetry (DSC). Calorimetric measurements showed that as the fusion protein increased, the main chain transition enthalpy decreased and the chain-melting temperature shifted, which is indicative of partial protein insertion into the hydrophobic region of the lipid membrane. This was confirmed using the Langmuir Blodgett technique (film balance) on lipid monolayers. The dissociation constant (K(d)) determined by the temperature jump method was approximately 1.1 microM.