Akinnuoye-Agbaje was born in Islington, London, to Nigerian parents of Yoruba origin, who were students in the UK. He is fluent in several languages, including Yoruba, Italian, and Swahili. He can "get along" in a few other languages, including French When he was only six weeks old, his biological parents gave him up to a white working-class family in Tilbury. His foster parents had at least ten African children, including Akinnuoye-Agbaje's two sisters, living in their house at certain points. His foster father made a living as a lorry driver and struggled to support the family financially.
Akinnuoye-Agbaje at a Red Cross benefit at the Hilton Hawaiian Village, 28 February 2007

When he was eight years old, his biological parents brought him back to Nigeria but, as he was unable to speak the Yoruba language and forbidden by his parents to speak English, he was returned to Tilbury shortly thereafter. The brief exposure to Nigeria left him struggling to reconcile his heritage with the distinctly English culture and environment he was raised in. As a teenager facing a cultural identity crisis, he joined a local skinhead gang in order to escape racial persecution at their hands. At 16 years old, having become a violent thief, his foster parents sent him to a boarding school in Surrey where he ultimately attempted suicide before coming to terms with his background and turning his life around.

He went on to earn a Law degree from King's College London and a Masters in Law from the University of London. While a university student, Akinnuoye-Agbaje worked in a clothes shop where he was introduced to the world of modeling. His modeling career eventually led him to Hollywood, where he began his acting career with a 1995 role in Congo.

His best known acting roles have been as the imposing convict Simon Adebisi in the 1990s HBO prison series Oz and as Mr. Eko on ABC's survivor drama Lost. He has numerous film credits since he began acting in 1994 and has appeared in many top films, including The Bourne Identity, in which he played a deposed African dictator, Hitu the police officer in Ace Ventura: When Nature Calls, Lock-Nah in The Mummy Returns, and Heavy Duty in G.I. Joe: The Rise of Cobra.

In 2009, Akinnuoye-Agbaje was in talks with Marvel Studios to play the superhero Black Panther in a proposed film of the same name. In an interview, he stated his excitement about the possibility, saying that "the timing is so right" for a black superhero, and "while I'm in my prime, this is the time... I'm going to keep knocking on their door." In 2014 Marvel did announce a Black Panther film, though with Chadwick Boseman in the title role.

Akinnuoye-Agbaje has also stated that he will be directing a film about his life story. More recently, he guest starred in the second episode of season 8 of Monk, and played Derek Jameson in the 2011 film The Thing. He portrayed Kurse in the Marvel Studios film Thor: The Dark World.