A unique nanostructure of Fe3O4 nanoparticles (NPs)-in/carbon layer/out-Ni NPs was developed and proved to be an efficient catalyst and protein adsorbent. This kind of nanostructure was formed through a space-confined pyrolysis procedure using polydopamine-Ni2+ coated Fe-NTA nanowires as the precursor. A N-doped carbon interlayer derived from polydopamine (PDA) supported a large amount of Ni NPs and entrapped well-defined Fe3O4 NPs, which were obtained through reduction of Ni2+, Fe3+ by carbonized NTA groups and a PDA layer. The contributions of the unique configuration along with the high density of Ni NPs in Fe3O4@C-Ni are significant for improving catalysis and protein adsorption performance, which is expected to be a promising alternative to other conventional catalysts and protein adsorbents. Due to the unique novel nanostructure, this nanocomposite possesses a wide range of applications, not only for catalytic reactions but also for other inhomogeneous reactions.