Protein–metabolite networks are central to biological systems, but are incompletely understood. Here, we report a screen to catalog protein–lipid interactions in yeast. We used arrays of 56 metabolites to measure lipid-binding fingerprints of 172 proteins, including 91 with predicted lipid-binding domains. We identified 530 protein–lipid associations, the majority of which are novel. To show the data set's biological value, we studied further several novel interactions with sphingolipids, a class of conserved bioactive lipids with an elusive mode of action. Integration of live-cell imaging suggests new cellular targets for these molecules, including several with pleckstrin homology (PH) domains. Validated interactions with Slm1, a regulator of actin polarization, show that PH domains can have unexpected lipid-binding specificities and can act as coincidence sensors for both phosphatidylinositol phosphates and phosphorylated sphingolipids ; 15 páginas, 8 figuras, 1 tabla ; This work is partially funded by Federal Ministry of Education and Research (BMBF) in the framework of the National Genome Research Network (NGFN) to ACG (BMBF NGNF IG-Cellular Systems Genomics, 01GS0865). OG is a fellow of the Ministerio de ciencia e innovación, Spain. KM is a fellow of the Danish Natural Science Research Council (09-064986/FNU). The protein interactions from this publication have been submitted to the IntAct database (pmid: 19850723) and assigned the identifier EBI-2933237