The dominant proteins released by Ascaris suum during development in vitro from the L3 to L4 stage were identified as collagenous cuticular proteins by sequence analysis and susceptibility to digestion by collagenase. Under reducing sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE), the collagen proteins separated into 3 groups with molecular weights estimated at 32 kDa, 54–60 kDa, and 71–91 kDa. The 32-kDa protein represents monomeric collagen; the 54–60- and 71–91-kDa components represent dimeric and trimeric forms, respectively, polymerized by nonreducible cross-links. Furthermore, the release of these forms of collagen was developmentally regulated, as exemplified by a sequential temporal progression from monomeric to dimeric to trimeric forms in association with the in vitro transition from L3 to L4. The data suggest that collagen released in vitro during development of A. suum L3 to L4 reflects the increased translation of collagen gene products and their initial assembly into higher molecular weight molecules associated with the synthesis of the L4 cuticle. A biotinylated dipeptidyl fluoromethylketone cysteine protease inhibitor (Bio-phe-ala-FMK) bound specifically to the 32-kDa collagen and, to a lesser extent, to a 30-kDa protein; binding was dependent on the presence of dithiothreitol (DTT) and was prevented by iodoacetamide. Because cysteine residues play an essential role in the initial assembly of the collagen monomers into the higher molecular weight oligomers present in the mature nematode cuticle, inhibition of molting of A. suum L3 to L4 by the cysteine protease inhibitor Z-phe-ala-FMK might be due to its binding to thiol groups of collagen monomers during a critical phase of collagen assembly. Prevention of cystine cross-links during this critical period of cuticle assembly by peptide-FMK inhibitors may represent a potential control mechanism having a novel mechanism of action.
CUTICULAR COLLAGEN SYNTHESIS BY ASCARIS SUUM DURING DEVELOPMENT FROM THE THIRD TO FOURTH LARVAL STAGE: IDENTIFICATION OF A POTENTIAL CHEMOTHERAPEUTIC AGENT WITH A NOVEL MECHANISM OF ACTION
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Marcia L. Rhoads, Raymond H. Fetterer, Joseph F. Urban, Jr.; CUTICULAR COLLAGEN SYNTHESIS BY ASCARIS SUUM DURING DEVELOPMENT FROM THE THIRD TO FOURTH LARVAL STAGE: IDENTIFICATION OF A POTENTIAL CHEMOTHERAPEUTIC AGENT WITH A NOVEL MECHANISM OF ACTION. J Parasitol 1 October 2001; 87 (5): 1144–1149. doi: https://doi.org/10.1645/0022-3395(2001)087[1144:CCSBAS]2.0.CO;2
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