The human mitochondrial proteome comprises approximately 1500 proteins, with only 13 being encoded by mitochondrial DNA. The remainder are encoded by the nuclear genome, translated by cytosolic ribosomes, and subsequently imported into and sorted within mitochondria. The process of mitochondria-destined protein import is mediated by several intricate protein complexes distributed among the four mitochondrial compartments. The focus of this mini-review is the translocase of the inner membrane 23 (TIM23) complex that assists in the import of ~60% of the mitochondrial proteome, which includes the majority of matrix proteins as well as some inner membrane and intermembrane space proteins. To date, numerous pathogenic mutations have been reported in the genes encoding various components of the TIM23 complex. These diseases exhibit mostly developmental and neurological defects at an early age. Interestingly, accumulating evidence supports the possibility that the gene for Tim50 represents a hotspot for disease-causing mutations among core TIM23 complex components, while genes for the mitochondrial Hsp70 protein (mortalin) and its J domain regulators represent hotspots for mutations affecting presequence translocase-associated motor (PAM) subunits. The potential mechanistic implications of the discovery of disease-causing mutations on the function of the TIM23 complex, in particular Tim50, are discussed.