We have analyzed the sequence of a mitochondrial integral membrane protein, Mdm12, and found that it forms the prototype for a novel domain, designated the SMP domain, that is common to an extended family of membrane-associated proteins. Comprehensive sequence searches using protein alignment models of SMP proteins were cross-validated by statistical resampling; providing strong support for these relationships. No consensus of 3-dimensional structure was reached upon threading sequences through known folds. SMP proteins are widespread amongst eukaryotic species with a particular enrichment in plants and features suggestive of species-specific functional variations. Members of 2 SMP families, the mitochore and tricalbin proteins, are essential components of protein complexes involved in mitochondrial inheritance and receptor endocytosis while a third SMP protein family, HT008, is associated with the Rvs161-Rvs167 complex, a known regulator of sphingolipid metabolism. In addition, HT008 and PDZK8 SMP proteins possess additional protein-protein interaction domains in domain architectures that are typical of protein scaffolds and adaptors. We therefore predict that the SMP domain is an important link between these distinct membrane-associated proteins and a key regulatory hub for unidentified global regulators.-Lee, I., Hong, W. Diverse membrane-associated proteins contain a novel SMP domain.