A cysteine protease from maize isolated in a complex with cystatin.
We recently purified a latent but SDS-activated protease complex (40, 15- or 13-kDa proteins) from maize [Yamada et al. (1998) Plant Cell Physiol. 39: 106]. Here, we revealed that the complex was composed of a cysteine protease (40 kDa) and a cystatin, cysteine protease inhibitor (15- or 13-kDa). This is the first report on the isolation of a complex consisting of a cystatin and a target cysteine protease from plants. Cloning of the cysteine protease revealed that it had low homology (25-30%) to other maize cysteine proteases cloned to date but was highly homologous to other plant cysteine proteases such as rice oryzain alpha (84%) and the homologs (50-80%). The cysteine protease expressed in Escherichia coli showed the same substrate and inhibitor specificities as the protease of the complex, demonstrating that the isolated cDNA clone exactly encodes the protease of the complex. The protease expressed in E. coli itself was active but not latent, probably because it was not bound to cystatin. It is most likely that in vitro activation of the protease complex by SDS is caused by the release of bound cystatin. The mRNA of protease was expressed in various tissues except for seeds.
Yamada
T
T
Department of Biological Sciences, Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, Japan. tyamada@bio.titech.ac.jp
Ohta
H
H
Shinohara
A
A
Iwamatsu
A
A
Shimada
H
H
Tsuchiya
T
T
Masuda
T
T
Takamiya
K
K
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D015891:Cystatins / Q000302:isolation & purification / Q000378:metabolism*; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D018744:DNA, Plant; D004789:Enzyme Activation; D004926:Escherichia coli; D015870:Gene Expression; D008969:Molecular Sequence Data; D010940:Plant Proteins; D012333:RNA, Messenger; D017386:Sequence Homology, Amino Acid; D012967:Sodium Dodecyl Sulfate; D003313:Zea mays / Q000201:enzymology*
D015891:Cystatins; D015853:Cysteine Proteinase Inhibitors; D018744:DNA, Plant; D010940:Plant Proteins; D012333:RNA, Messenger; D012967:Sodium Dodecyl Sulfate; C411583:CPPIC protein, Zea mays; D003546:Cysteine Endopeptidases