Localization of Phenol Oxidase in Female Trichuris suis

Abstract

A phenol oxidase (E.C. 1.10.3.1) preparation from adult female Trichuris suis was assayed by both polarographic and spectrophotometric techniques. The T. suis enzyme oxidized most diphenols including 4-methylcatechol (4MC) and dihydroxyphenylalanine but did not oxidize tyrosine. The pH and temperature optima were 6.8 and 36 C, respectively. The Km measured using 4MC as a substrate ranged from 0.12 to 0.4 mM. The highest phenol oxidase activity was isolated in fractions from the adult females that were enriched in eggs relative to the activity in somatic tissue from females and all male tissues that were assayed. Phenol oxidase activity was localized on sodium dodecyl sulfate-polyacrylamide gel electrophoresis substrate gels into 2 bands with M(r)'s of 44,000 and 53,000. An antibody to the 44,000 band recognized 2 bands of 40,000 and 45,000 M(r) on western blot analysis of the enzyme preparation. Immunocytochemical localization of anti-phenol oxidase antibody in serial cross sections of adult female worms indicates that the enzyme is found exclusively in the anterior part of the parasite in the proximal part of the uterus that is posterior to the junction with the stichosome. Eggs located in more distal parts of the reproductive system did not react with the antibody. The results indicate that a phenol oxidase is located in the fertilized eggs of adult female T. suis. It is likely that phenol oxidase contributes significantly to the chemical hardening process in the eggs when they pass out into the external environment. Inhibition of phenol oxidase may reduce the survivability of the eggs and thus minimize contamination of livestock facilities.