The emergence of Middle East respiratory syndrome coronavirus (MERS-CoV), a deadly human coronavirus, has triggered considerable interest in the biomedical community. Similar to other enveloped viruses, coronaviruses access host cells by membrane fusion, a process mediated by specific fusion or “spike” proteins on the virion, often activated by cellular proteases. We have identified unique features of the MERS-CoV spike (S) protein cleavage activation. Our findings suggest that S can be activated by furin, a broadly expressed protease, by a two-step cleavage mechanism, occurring at distinct sites, with cleavage events temporally separated. Such furin-mediated activation is unusual in that it occurs in part during virus entry. Our findings may explain the polytropic nature, pathogenicity, and life cycle of this zoonotic coronavirus.