Function of the N-terminal propeptide of an aminopeptidase from Vibrio proteolyticus.
An aminopeptidase from Vibrio proteolyticus was translated as a preproprotein consisting of four domains: a signal peptide, an N-terminal propeptide, a mature region and a C-terminal propeptide. Protein expression and analysis of the activity results demonstrated that the N-terminal propeptide was essential to the formation of the active enzyme in Escherichia coli. Urea dissolution of inclusion bodies and dialysis indicated that the N-terminal propeptide could facilitate the correct folding of the enzyme in vitro. Using L-Leu-p-nitroanilide as the substrate, the kinetic parameters (k(cat) and K(m)) of the pro-aminopeptidase and processed aminopeptidases were analysed. The results suggested that the N-terminal propeptide inhibited enzyme activity of the mature region. In contrast, the C-terminal propeptide did not show evidence of forming an active enzyme, of correctly folding in vitro or of inhibiting the active region.
Zhang
Z Z
ZZ
Applied Enzymology Laboratory, National Food Research Institute, Tsukuba, Ibaraki 305-8642, Japan.
Nirasawa
S
S
Nakajima
Y
Y
Yoshida
M
M
Hayashi
K
K
D000626:Aminopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D011489:Protein Denaturation; D020673:Protein Renaturation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014733:Vibrio / Q000201:enzymology*
D017931:DNA Primers; D011994:Recombinant Proteins; D000626:Aminopeptidases