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Requirement for the COOH-terminal pro-sequence in the translocation of aqualysin I across the cytoplasmic membrane in Escherichia coli.

 Dong-Wook Kim  ;  Hiroshi Matsuzawa 
 Biochemical and Biophysical Research Communications, Vol.277(1) : 216-220, 2000 
Journal Title
 Biochemical and Biophysical Research Communications 
Issue Date
Cell Membrane/metabolism* ; Enzyme Activation ; Escherichia coli/cytology ; Escherichia coli/metabolism* ; Mutation/genetics ; Protein Conformation ; Protein Precursors/chemistry ; Protein Precursors/genetics ; Protein Precursors/metabolism ; Protein Processing, Post-Translational ; Protein Sorting Signals/genetics ; Protein Sorting Signals/physiology* ; Protein Transport ; Recombinant Proteins/chemistry ; Recombinant Proteins/metabolism ; Serine Endopeptidases/chemistry ; Serine Endopeptidases/genetics ; Serine Endopeptidases/metabolism* ; Thermus/enzymology*
aqualysin I ; serine protease ; C-terminal pro-sequence ; protein secretion ; protein translocation ; intramolecular chaperone
Aqualysin I from Thermus aquaticus YT-1 is an extracellular subtilisin-type serine protease. The protease is synthesized as a distinct precursor composed of four functional domains: an N-terminal signal sequence, an N-terminal pro-sequence, a protease domain, and a C-terminal pro-sequence. The N-terminal pro-sequence is essential for the production of active aqualysin I while the C-terminal pro-sequence is required for extracellular secretion of aqualysin I. In an E. coli expression system, the function of C-terminal pro-sequence in the translocation of aqualysin I across the cytoplasmic membrane was investigated. More than 60-70% of the total activity was detected in the cytoplasmic fraction in the deletion mutations of the C-terminal pro-sequence while less than 30% was found in this fraction in wild type. In addition, in vitro processing of aqualysin I precursors with these mutations to a mature form promptly occurred and the folding into active aqualysin I was rapid. These results suggest that the C-terminal pro-sequence, probably in conjunction with the signal sequence, facilitates the translocation of the precursor across the cytoplasmic membrane by preventing the precursor from taking on an active conformation.
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1. College of Medicine (의과대학) > Dept. of Physiology (생리학교실) > 1. Journal Papers
Yonsei Authors
Kim, Dong Wook(김동욱) ORCID logo https://orcid.org/0000-0002-5025-1532
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