Autolytic activation mechanism of Bombyx acid cysteine protease (BCP).
Acid cysteine proteinase in the eggs of the silkmoth, Bombyx mori, exists as an inactive proenzyme. This 47-kDa pro-BCP1 zymogen molecule can be processed in vitro into an enzymatically active 39-kDa BCP molecule. In this current study, the maximum rate of processing in vitro was achieved at approximately pH 4.0, at a temperature of 37 degrees C under reducing conditions. The rate of conversion was not affected by increasing concentrations of pro-BCP. We prepared immobilized BCP bound to AH-Sepharose and examined the activation. Immobilized pro-BCP was autolysed, although the rate of processing was slow, indicating that the reaction might be an intramolecular one. Kinetic experiments suggest that the mechanism is likely to involve a stepwise reaction, in which pro-BCP is converted to an active enzyme through intermediate forms releasing small peptides stepwise. The results suggest that autocatalytic cleavage (intramolecular) is a major processing step in the early stage of pro-BCP activation.
Takahashi
S Y
SY
Department of Biochemistry and Radiation Biology, College of Agriculture, Yamaguchi University, Japan.
Yamamoto
Y
Y
Watabe
S
S
Kageyama
T
T
D000595:Amino Acid Sequence; D000818:Animals; D012831:Bombyx / Q000201:enzymology*; D002384:Catalysis; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D004527:Egg Proteins / Q000737:chemistry / Q000378:metabolism*; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000378:metabolism*; D004800:Enzymes, Immobilized; D006863:Hydrogen-Ion Concentration; D019476:Insect Proteins / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013696:Temperature
D004527:Egg Proteins; D004792:Enzyme Precursors; D004800:Enzymes, Immobilized; D019476:Insect Proteins; D010446:Peptide Fragments; D003546:Cysteine Endopeptidases; C108394:acid cysteine proteinase, Bombyx