Zn-binding protein was obtained after purification from scallops (Mizuhopecten yessoensis) using gel permeation and ion-exchange chromatography. Amino acid determination result shows that the cysteine of the zinc-binding protein accounts for one third of the total amino acids, which is a typical feature of metallothionein (MT). The secondary structure of zinc-binding protein was determined by Fourier Transform Infrared Spectroscopy (FTIR) and Circular dichroism (CD), the result shows that the α-helix was 46.55%, the β-sheets was 27.07%, the random coil was 16.48% and the β-turns was 9.89%. Using a commercial kit to measure its in vitro antioxidant activity, the result shows that it has good scavenging ability to 1, 1-diphenyl-2-picrylhydrazyl (DPPH), Hydroxyl radical (•OH) and reducing ability to ferrous iron ions. In this study, the extraction and purification of zinc-binding protein can be prepared in large quantities, which provide a basis for its industrialization.