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The translocase of the inner membrane 22 (TIM22) is a multisubunit molecular machine specialized for the translocation of hydrophobic, multi\u2010transmembrane\u2010spanning proteins with internal targeting signals into the inner mitochondrial membrane. Here, we undertook a crosslinking\u2010mass spectrometry (XL\u2010MS) approach to determine the molecular arrangement of subunits of the human TIM22 complex. Crosslinking of the isolated TIM22 complex using the BS3 crosslinker resulted in the broad generation of crosslinks across the majority of TIM22 components, including the small TIM chaperone complex. 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