Penghlis starred in the 1980's revival of Mission: Impossible, which was filmed in his native Australia. Penghlis' role as actor, makeup Artist, and voice impersonator Nicholas Black in the revival was a counterpart to Martin Landau's Rollin Hand and Leonard Nimoy's The Great Paris.

In 2003, Penghlis was nominated for the Soap Opera Digest Award for Favorite Return for his return to the cast of Days of our Lives. He left the show in fall 2005. Penghlis returned to Days on 24 May 2007 to reprise the role of André DiMera. He also began reprising the role of nice guy Tony DiMera on 17 July 2007. Penghlis was nominated for Outstanding Leading Actor at the Daytime Emmy Awards in 2008. In 2009, Penghlis' character was once again written out by having him die. In 2010 Penghlis returned to stage acting, performing in New Jersey's Cape May Stage theatre. He starred in Charles Evered's play Class with Heather Matarazzo.

Penghlis played a role in the 2013 film The Book of Esther.

Penghlis was born in Surry Hills, New South Wales of Greek parentage. Penghlis started his career in acting on stage, performing in Jockeys under Milton Katselas's direction. Penghlis first appeared to daytime audiences in 1981 on General Hospital during the Ice Princess saga when he played the role of Victor Cassadine, brother of Mikkos and Anthony Cassadine. After his character was written out of the show by being taken to prison, he was cast as the villainous Count Tony DiMera, the son of villain Stefano DiMera in the NBC daytime drama Days of Our Lives. Penghlis returned to daytime in his one-time role of Victor Cassadine on General Hospital from 30 January 2014 to 4 March 2014.