The light-driven proton pump Coccomyxa subellipsoidea rhodopsin (CsR) provides—because of its high expression in heterologous host cells—an opportunity to study active proton transport under controlled electrochemical conditions. In this study, solving crystal structure of CsR at 2.0-Å resolution enabled us to identify distinct features of the membrane protein that determine ion transport directivity and voltage sensitivity. A specific hydrogen bond between the highly conserved Arg⁸³ and the nearby nonconserved tyrosine (Tyr¹⁴) guided our structure-based transformation of CsR into an operational light-gated proton channel (CySeR) that could potentially be used in optogenetic assays. Time-resolved electrophysiological and spectroscopic measurements distinguished pump currents from channel currents in a single protein and emphasized the necessity of Arg⁸³ mobility in CsR as a dynamic extracellular barrier to prevent passive conductance. Our findings reveal that molecular constraints that distinguish pump from channel currents are structurally more confined than was generally expected. This knowledge might enable the structure-based design of novel optogenetic tools, which derive from microbial pumps and are therefore ion specific.