We report the preparation of spherical protein nanoparticles comprising self-assembled human serum albumin (HSA) with an N-(11-carboxyundecyl)succinimido (CU) arm in aqueous medium at pH 5.0. Molecular dynamics simulation implied that the alternate CU chain bindings and multiple hydrogen bonds contribute the interprotein interaction. Cross-linking of the protein components by glutaraldehyde stabilized the particle under physiological pH.