Levandowski is at the center of Uber's intense legal battle with Waymo, the self-driving car unit of Google's parent Alphabet. In Waymo's suit, the company accuses Levandowski, a former Google engineer, of stealing 14,000 "highly confidential" files on driverless car technology.
Instead of cooperating with the suit, Levandowski pleaded the Fifth Amendment, which protects individuals from self-incrimination.
An Uber spokeswoman confirmed the termination. The New York Times earlier reported news of the firing.
Uber had urged Levandowski to cooperate with an internal investigation for months, said the spokeswoman on Tuesday. Uber set a deadline for him to comply, but he didn't meet it.
The spokeswoman added that people who reported to Levandowski will now report to Eric Meyhofer, who took charge of Uber's Advanced Technologies Group in April.
The lawsuit pits two high-profile Silicon Valley companies against each other. What's at stake is the future of self-driving car technology, which could profoundly change transportation. Google has been working on self-driving cars since 2009, and Uber has been betting on self-driving cars as the next big thing for its company.
Earlier this month, Uber made clear that it wasn't messing around when it came to Levandowski's cooperation with the investigation. "If you fail to comply in a material manner," Sally Yoo, Uber's head of legal, wrote in a letter to the engineer, "then Uber will take adverse employment action against you, which may include termination of your employment."
Waymo didn't immediately respond to a request for comment on Levandowski's termination.
Special Reports: CNET's in-depth features in one place.
Technically Literate: Original works of short fiction with unique perspectives on tech, exclusively on CNET.