The lawsuit, filed Tuesday in federal court, asks a judge to stop state officials from preventing gay couples from getting married through the state's current version of the Defense of Marriage Act.
The lawsuit names Kane and Governor Tom Corbett, a Republican who opposes gay marriage, among the defendants.
But speaking at the National Constitution Center in Philadelphia, Kane said she "cannot defend" state law that prohibits same-sex marriage. She also said she had not spoken with the governor about her decision.
Although state law says the attorney general is supposed to defend the constitutionality of Commonwealth laws, the attorney general may allow lawyers for the governor's office or other executive agencies to defend a suit.
She said "ripples can create a wave to break down walls of intolerance," winning loud applause for the Robert F. Kennedy quote.
"We are the land of the free and the home of the brave, and I want to start acting like that," said Kane, who said on the campaign trail last year that gay people should be allowed to marry.
State Rep. Brian Sims, a civil rights attorney and Pennsylvania's first openly LGBT elected state official, posted on Twitter Thursday morning, "Attorney General Kathleen Kane, if you were a man I'd marry you!"
Legal experts believe the case is bound for the U.S. Supreme Court, along with similar bans in other states, forcing a decision on whether it is unconstitutional to deny same-sex couples the opportunity to marry.