Recognition and processing of a nuclear-encoded polyprotein precursor by mitochondrial processing peptidase.
The nuclear-encoded protein RPS14 (ribosomal protein S14) of rice mitochondria is synthesized in the cytosol as a polyprotein consisting of a large N-terminal domain comprising preSDHB (succinate dehydrogenase B precursor) and the C-terminal RPS14. After the preSDHB-RPS14 polyprotein is transported into the mitochondrial matrix, the protein is processed into three peptides: the N-terminal prepeptide, the SDHB domain and the C-terminal mature RPS14. Here we report that the general MPP (mitochondrial processing peptidase) plays an essential role in processing of the polyprotein. Purified yeast MPP cleaved both the N-terminal presequence and the connector region between SDHB and RPS14. Moreover, the connector region was processed more rapidly than the presequence. When the site of cleavage between SDHB and RPS14 was determined, it was located in an MPP processing motif that has also been shown to be present in the N-terminal presequence. Mutational analyses around the cleavage site in the connector region suggested that MPP interacts with multiple sites in the region, possibly in a similar manner to the interaction with the N-terminal presequence. In addition, MPP preferentially recognized the unfolded structure of preSDHB-RPS14. In mitochondria, MPP may recognize the stretched polyprotein during passage of the precursor through the translocational apparatus in the inner membrane, and cleave the connecting region between the SDHB and RPS14 domains even before processing of the presequence.
Department of Chemistry, Faculty of Science, Kyushu University, Fukuoka 812-8581, Japan.
Tsutomu
T
Oshima
Eiki
E
Yamasaki
Tadashi
T
Ogishima
Koh-ichi
K
Kadowaki
Akio
A
Ito
Sakae
S
Kitada
D002467:Cell Nucleus / Q000235:genetics*; D008666:Metalloendopeptidases / Q000235:genetics / Q000378:metabolism*; D008928:Mitochondria / Q000201:enzymology* / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D012275:Oryza; D020815:Polyproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017510:Protein Folding; D011498:Protein Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity; D013385:Succinate Dehydrogenase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000096122:Mitochondrial Processing Peptidase
D008666:Metalloendopeptidases; D020815:Polyproteins; D011498:Protein Precursors; D012269:Ribosomal Proteins; D013385:Succinate Dehydrogenase; D000096122:Mitochondrial Processing Peptidase; C039747:ribosomal protein S14