Characterization of novel cysteine proteases from germinating cotyledons of soybean [Glycine max (L.) Merrill].
The enzymatic properties of novel cysteine proteases D3-alpha and beta which were purified from germinating soybean cotyledons were investigated. The enzyme activities were exhibited in the presence of a thiol reagent, such as 2-mercaptoethanol, and apparently inhibited by E-64, a cysteine protease inhibitor. Hydrolytic activities toward carbobenzoxy-Phe-Arg-MCA were detected at a pH above 4.0. The optimum temperature for activities was about 40 degrees C. The isoelectric point of D3-alpha and beta was 4.4 and 4. 7, respectively. The molecular mass of D3-alpha and beta, measured by MALDI/TOF mass spectrometry, was 26,178 and 26,429 Da, respectively. The substrate specificities of the enzymes were examined using peptide-MCAs and peptides, and cathepsin L-like broad specificity was observed at pH 4.0. These results demonstrated that these enzymes are cysteine endopeptidases [EC 3.4.22.-] like papain [EC 3.4.22.2].
Asano
M
M
Food Research & Development Laboratories, Ajinomoto Co., Inc., Kawasaki-ku, Kawasaki, Kanagawa, 210-8681, Japan. fld_asano@te11.ajinomoto.co.jp
Suzuki
S
S
Kawai
M
M
Miwa
T
T
Shibai
H
H
D000595:Amino Acid Sequence; D002845:Chromatography; D003546:Cysteine Endopeptidases / Q000737:chemistry* / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D005934:Glucagon / Q000494:pharmacology; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000737:chemistry; D008969:Molecular Sequence Data; D009496:Neurotensin / Q000494:pharmacology; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification; D011480:Protease Inhibitors / Q000494:pharmacology; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D013025:Glycine max / Q000201:enzymology*; D013696:Temperature
D007527:Isoenzymes; D010940:Plant Proteins; D011480:Protease Inhibitors; D011500:Protein Synthesis Inhibitors; D007987:Gonadotropin-Releasing Hormone; D009496:Neurotensin; D005934:Glucagon; D003546:Cysteine Endopeptidases