title: Transepithelially transported pro-phenoloxidase in the cuticle of the silkworm, Bombyx mori. Identification of its methionyl residues oxidized to methionine sulfoxides.
abstract: Pro-phenoloxidase (proPO) in insects is activated through the action of a protease cascade triggered by minute amounts of microbial cell wall components. It is an important molecule for the defense against invading microorganisms and for the repair of wounds. In the accompanying paper (Asano, T., and Ashida, M. (2001) J. Biol. Chem. 276, 11100-11112), a proPO isoform, proPO-HS, in the hemolymph of the silkworm, Bombyx mori, is reported to be transported to the cuticle. The transported proPO isoform was recovered from the cuticle and named proPO-CS. The elution profiles of proPO-CS and proPO-HS in reversed-phase high performance liquid chromatography (HPLC) were found to be different, giving a basis to the inference that proPO-CS is a modified form of proPO-HS. In the present study, we investigated the nature of the modifications occurring in proPO-CS, in which proteolytically and chemically cleaved fragments originating from the subunits of proPO-CS and proPO-HS were analyzed by reversed-phase HPLC, amino acid sequencing, and mass spectrometry. A subunit of the heterodimeric proPO-CS was found to contain five or six methionine sulfoxides, and another subunit was found to contain one methionine residue oxidized to the sulfoxide. All of the oxidized methionyl residues were identified. Other than oxidation of the methionyl residues, no additional modification of proPO-CS was found. In the model structure of each subunit of proPO-CS constructed by protein modeling with the known structures of the horseshoe crab, Limulus polyphemus, hemocyanin type II subunit as templates, the methionine residues identified as methionine sulfoxide had high degrees of accessibility to the solvent. The implication of the oxidation at the methionine residues is discussed in relation to the mechanism of transepithelial transport of proPO from the hemolymph to the cuticle.
authors: Biochemistry Laboratory, The Institute of Low Temperature Science, Hokkaido University, Sapporo 060-0819, Japan.
authors: T
authors: T
authors: Asano
authors: M
authors: M
authors: Ashida
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D012831:Bombyx / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D008715:Methionine / Q000031:analogs & derivatives* / Q000378:metabolism*; D008969:Molecular Sequence Data; D014442:Monophenol Monooxygenase / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D010802:Phylogeny; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D012867:Skin / Q000378:metabolism*
chemical_list: D011498:Protein Precursors; D008715:Methionine; D014442:Monophenol Monooxygenase; C013111:methionine sulfoxide