Characterizing Complexation of Charge Sequence Specific Atactic Polyampholyte Peptides

Polyampholytes (PAs) are polymers containing both positively and negatively ionizable groups along their backbone. Intrinsically disordered proteins (IDPs) are one of the most prevalent examples of PA systems. To gain a greater physical understanding of electrostatic contribution to PA conformation and IDP structures, we designed a set of 5 atactic, charge sequence varied block copolymer peptides to experimentally observe and evaluate electrostatic interactions.

Using solid phase synthesis, we synthesized these PA peptides using 16 randomly chiral (D,L)-glutamic acid (anionic) and 16 randomly chiral (D,L)-lysine (cationic) residues arranged in a charged blocks of 1 (alternating sequence) to 16 residues (diblock sequence).

From FTIR and CD measurements, these atactic PA peptides show no secondary structure formation, allowing us to focus on charge sequence effects on electrostatic interactions. We also observe sequence-dependent liquid-liquid phase separation from turbidity measurements. Interestingly, from SAXS experiments, these PAs begin forming multi-chain complexes with larger block sizes. However, quantifying the size and shape of these complexes is challenging as there are few models which can capture both single chain and multi-chain complexes in solutions. To overcome this challenge, we apply the Pedersen-Schurtenberger semi-flexible polymer with excluded volume model form factor with polydisperse contour length to capture and quantify the complexation behavior of these atactic PA peptide solutions. From our analysis of PA solutions' SAXS profiles, we observe increasing complexation with increasing charge block size. We also characterize the dependence of complexation on PA peptide concentration and NaCl concentration for each block size. These results shed light onto how charge sequence influences the thermodynamics of PA and IDP systems leading to a wide range of single and multi-chain conformations.