Structural and enzymatic characterization of physarolisin (formerly physaropepsin) proves that it is a unique serine-carboxyl proteinase.
Previously, we purified and partially characterized physarolisin, a lysosomal acid proteinase from Physarum polycephalum, which had been suggested to be concerned with the morphological changes of the mold. In this study, a cDNA for the enzyme was cloned and sequenced, and the structural and enzymatic features were investigated. The enzyme shows a sequence similarity to the serine-carboxyl proteinase family (MEROPS S53). Indeed, diisopropylfluorophosphate (DFP) was shown to strongly inhibit the activity of the enzyme. However, the enzyme possesses several unique features distinct from the other members of the family, such as the two-chain structure and inhibition by diazoacetyl-D,L-norleucine methyl ester (DAN). The sites and mode of processing of the precursor to the mature enzyme were deduced, and the major DAN-reactive residue in the enzyme was identified to be Asp529. These features were suggested to be due to the unique local tertiary structure of the enzyme by molecular modeling. We now propose the name physarolisin for the enzyme.
Nishii
Wataru
W
School of Life Science, Tokyo University of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, 192-0392, Tokyo, Japan.
Ueki
Tomoko
T
Miyashita
Rie
R
Kojima
Masaki
M
Kim
Yong-Tae
YT
Sasaki
Narie
N
Murakami-Murofushi
Kimiko
K
Takahashi
Kenji
K
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D020134:Catalytic Domain; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D016054:DNA, Protozoan / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009646:Norleucine / Q000031:analogs & derivatives*; D016839:Physarum polycephalum / Q000201:enzymology / Q000235:genetics; D011487:Protein Conformation; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D018076:DNA, Complementary; D016054:DNA, Protozoan; D015800:Protozoan Proteins; C000909:N-diazoacetylnorleucine methyl ester; D009646:Norleucine; D012697:Serine Endopeptidases