Inspired by the molecular mechanics of mussel adhesive formation, a novel water-soluble fluorescent macromolecule (polydopamine-polyethyleneimine (PDA-PEI)) is prepared by one-pot copolymerization of dopamine (DA) and PEI. In this method, DA is polymerized to form PDA, which is then coupled with PEI mainly through Michael addition. The fluorescence property of PDA-PEI is mainly attributed to the Michael addition of PEI on the 5,6-dihydroxyindole (DHI) units of PDA, where PEI can form hydrogen bonds with oxidative products such as DHI and force the DHI units to twist out of plane, resulting in a decrease in the intra- and intermolecular coupling of PDA. In addition, the influence of various metal cations on the fluorescence of the PDA-PEI copolymer is investigated. This work may facilitate the development of new strategies for controlling the emission characteristics of PDA.