In Saccharomyces cerevisiae, the heteromeric HAP transcription factor is necessary for optimal growth on respiratory carbon sources. One of its components, the Hap4p protein, is necessary for transcriptional activation. The same protein is also the regulatory part of the complex in response to carbon sources, as HAP4 is strongly induced during the shift from fermentative to respiratory metabolism in S. cerevisiae. We report here the characterization of a new gene from the respiratory yeast Kluyveromyces lactis, obtained by heterologous complementation of a delta hap4 S. cerevisiae mutant strain. The deduced sequence of the protein (643 amino acids) exhibits two small domains (11 and 16 amino acids respectively) highly homologous to corresponding domains of ScHap4p, while the overall similarity is rather weak. Additional experiments were performed to confirm the functional homology of this new gene with ScHAP4, which we named KIHAP4. The importance of the small highly conserved N-terminal sequence was confirmed by in vitro mutagenesis. All the mutations that interfere with the Hap4p-Hap2/3/5 interaction were localized in it. The discovery of the same regulatory protein in two metabolically distinct yeast species raises the question of its functional significance during evolution.