The biogenesis of mitochondria relies on the import of hundreds of different precursor proteins from the cytosol. Most of these proteins are synthesized with N-terminal presequences which serve as matrix targeting signals. Presequences consistently form amphipathic helices, but they considerably differ in respect to their primary sequence and length. Here we show that presequences can be grouped into different classes on basis of their specific features. Using a test set of different presequences, we observed that Class 1 presequences endow precursor proteins with improved in vitro import characteristics. We developed IQ-compete, a novel fluorescence de-quenching-based assay to monitor the import efficiencies of mitochondrial precursors in vivo. With this assay, we confirmed the increased import competence of Class 1 presequences. Using mass spectrometry, we found that the presequence of the Class 1 protein Oxa1 specifically recruits the tetratricopeptide repeat (TPR) containing protein TOMM34 to the cytosolic precursor protein. TOMM34 and its yeast homolog Cns1 apparently serve as presequence-specific targeting factors that increase the import efficiency of a specific subset of mitochondrial precursor proteins. Our results suggest that presequences contain a protein-specific priority code that encrypts the targeting mechanism of individual mitochondrial precursor proteins.