title: Purification and characterization of a cysteine proteinase from silkworm eggs.
abstract: Eggs of the silkworm, Bombyx mori, contain a high level of a proteinase which is most active in acidic pH region. The proteinase was purified from an extract of eggs by a six-step procedure which included conventional chromatographic fractionations. The molecular mass of the proteinase was estimated to be 350 kDa by gel filtration and 47 kDa by electrophoresis on sodium dodecyl sulfate/polyacrylamide gels, suggesting an octameric structure. The amino acid composition was found to resemble that of mammalian lysosomal cysteine proteinases, in particular cathepsin L. The NH2-terminal 10-residue sequence is Val-Gln-Phe-Phe-Asp-Leu-Val-Lys-Glu-Glu-. The enzyme appears to be a member of the class of cysteine proteinases since it was strongly inhibited by sulfhydryl-reactive compounds and N-[N-(1,3-trans-carboxyoxiran-2-carbonyl)-L-leucyl]-agmatine (E-64). The enzyme hydrolyzed various protein substrates, such as hemoglobin, vitellogenin, vitellin, and lipophorin, with maximal activity around pH 3-3.5. The specificity of the cleavage sites in the oxidized B chain of insulin was rather well defined and there was high affinity for hydrophobic residues at the P2 and P3 positions. The cysteine proteinase is thought to be involved in protein degradation during embryonic development of silkworm eggs.
authors.affiliation: Department of Biochemistry, Kyoto University, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Kageyama
authors.forename: S Y
authors.initials: SY
authors.lastname: Takahashi
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D012831:Bombyx / Q000201:enzymology*; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification*; D015853:Cysteine Proteinase Inhibitors; D004527:Egg Proteins / Q000378:metabolism; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010063:Ovum / Q000201:enzymology*; D013379:Substrate Specificity
chemical_list: D000596:Amino Acids; D015853:Cysteine Proteinase Inhibitors; D004527:Egg Proteins; D003546:Cysteine Endopeptidases