The duo, who sold millions of albums and enjoyed 1980s chart successes with records such as Sweet Dreams [Are Made of This] and There Must be an Angel, will take part in a 50th anniversary celebration of The Beatles' appearance on US TV's Ed Sullivan Show.
It comes seven years after Lennox, from Aberdeen, said she didn't foresee them working together again. They last toured together in 2000, following the release of their album Peace and have since concentrated on solo work.
They will play songs by The Beatles at the gig on January 27, which is organised by the Recording Academy, which hands out the annual Grammy Awards.
The Eurythmics will also share the stage with R&B singers Alicia Keys and John Legend and pop-rock group Maroon 5.
The Night that Changed America: A Grammy Salute to The Beatles pays homage to the Fab Four's February 1964 performances on the CBS show.
Ken Ehrlich, who is also behind the Beatles special, said: "When it came to booking this show, what I felt was important was to try and find those artists who not only would be able to interpret Beatles songs, but would also have an... understanding what they meant."