A member of the Wayans family of entertainers, Damon performed as a comedian and actor throughout the 1980s, including a brief stint on the NBC sketch comedy series Saturday Night Live.

Since then, he has starred in a number of films and television shows, some of which he has co-produced or co-written, including Mo' Money, The Last Boy Scout, Major Payne, Bulletproof, and the sitcoms My Wife and Kids and Poppa's House, the latter featuring his son Damon Wayans Jr. From 2016 to 2019, he starred as Roger Murtaugh in the Fox television series Lethal Weapon.

With his brother Keenen, Wayans created the Fox sketch comedy series In Living Color, which had a mostly African-American cast.

In October 1996, he produced Waynehead, a short-lived cartoon for The WB, loosely based on his own childhood growing up in a large family, starring a poor boy with a club foot.

From 1997 to 1998, he was the executive producer of 413 Hope St., a short-lived drama on the FOX network starring Richard Roundtree and Jesse L. Martin.

In 1999, his The New York Times bestselling book Bootleg, with co author David Asbery was published; it is a humorous compilation of his observations about family.

In the end of 2006, he produced and starred in the Showtime sketch comedy series The Underground, which also featured his son, Damon Jr.

As of 2014[update], Wayans continues to perform stand-up comedy and has developed apps with his company of freelancers "MIMS" (Money in My Sleep).

In September 2016, he was cast as Roger Murtaugh in the television version of Lethal Weapon, a role originated by Danny Glover in the film series.

Wayans returned to scripted television in the fall of 2024 with his son Damon Jr. on the CBS sitcom Poppa's House.