FIGURE 3:

Deletion of MRX15 together with MBA1 leads to respiratory deficiency and altered membrane attachment. (A) Serial-dilution growth test on full medium fermentable (glucose) and nonfermentable (glycerol) carbon sources of indicated strains. In the mrx15∆C mutant, the terminal 126 amino acids were replaced by a PA tag. In the C-terminal Oxa1 mutant the 71 C-terminal residues were deleted alone or together with MRX15. Cells were grown to logarithmic phase, spotted in 10-fold dilutions, and incubated at 30 and 37°C. (B) Ribosome pelletation at different ionic strengths. The ribosome and attached factors were separated after lysis in different salt conditions by a sucrose cushion and high-velocity centrifugation. The different fractions were probed for a soluble protein (Aco1), a ribosomal marker (Mrpl36), Mba1, and PA for Mrx15 detection. (C) Flotation gradient of mitochondrial extracts from wild-type or mrx15∆mba1∆ strains. Soluble and membrane fractions were separated after freeze–thaw cycles by a sucrose step gradient and high-velocity centrifugation. Fractions were probed for the LSU (Mrpl36, Mrpl4, and Mrpl40), membrane (Cbp4), and soluble proteins (Aco1). P, pellet; SN, supernatant; T, total; WT, wild type.