Synthesis of tyrosine-derived cross-links in Ascaris suum cuticular proteins.
Tritiated dityrosine and isotrityrosine were detected by high performance liquid chromatography (HPLC) of acid hydrolysates of cuticular proteins from larval Ascaris suum following their 96-hr in vitro incubation in [3H]tyrosine. Sixty percent of the HPLC-recovered radiolabel was present as tyrosine, 20% as dityrosine, and 6% as isotrityrosine. Approximately 13% of radioactivity was associated with several unidentified peaks. A similar distribution of radioactivity was observed in acid hydrolysates of cuticular proteins from young adults of A. suum following 48 hr in vitro incubation with [3H]tyrosine. The 2-mercaptoethanol (2ME)-insoluble cuticular protein from the larval stages had a higher rate of synthesis of [3H]dityrosine than did the 2ME-soluble cuticular proteins, whereas the 2ME-soluble cuticular proteins had higher rates of synthesis of [3H]isotrityrosine. Pulse-chase studies of A. suum larvae demonstrated a relatively low rate of synthesis of both dityrosine and isotrityrosine. The addition to the culture media of the peroxidase inhibitors, phenylhydrazine (PHEN), 3-amino-1,2,4-triazole (AT), and N-acetyltyrosine (NAT) reduced the amount of [3H]tyrosine synthesized into both dityrosine and isotrityrosine. In a cell-free system, soluble extracts of A. suum larvae also converted radiolabeled tyrosine to dityrosine; isotrityrosine was produced by some extracts. The rate of conversion correlated with time of incubation and the volume of added extract and was inhibited by AT, NAT, and PHEN, with PHEN being the most potent inhibitor. The results of the present study suggest that the tyrosine residues of the cuticular proteins are posttranslationally modified by the formation of dityrosine and isotrityrosine cross-links. This modification is most likely mediated by a peroxidase.
Fetterer
R H
RH
Helminthic Diseases Laboratory, Agricultural Research Service, U.S. Department of Agriculture, Beltsville, Maryland 20705.
Rhoads
M L
ML
Urban
J F
JF
D000818:Animals; D017165:Ascaris suum / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D007814:Larva / Q000378:metabolism; D013552:Swine; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000032:analysis / Q000096:biosynthesis
D014443:Tyrosine; C030231:isotrityrosine; C007543:dityrosine