It happened during an outburst by Bryant after he received his 15th technical foul of the season.
Game video shows an upset Kobe hit a chair, throw a towel and then yell at referee Bennie Adams, calling him a "f***ing f**got."
Bryant responded Wednesday morning after growing criticism about the remark.
"What I said last night should not be taken literally," Bryant said in a statement. "My actions were out of frustration during the heat of the game. Period. The words expressed do not reflect my feelings towards the gay and lesbian communities and were not meant to offend anyone."
In announcing the fine, NBA issued a statement from Commissioner David Stern.
"Kobe Bryant's comment during last night's game was offensive and inexcusable," Stern said. "While I'm fully aware that basketball is an emotional game, such a distasteful term should never be tolerated. Kobe and everyone associated with the NBA know that insensitive or derogatory comments are not acceptable and have no place in our game or society."
One of TNT's announcers, Steve Kerr, noticed, and immediately remarked, "Might want to take the camera off him right now, for the children watching."
The Lakers star could receive a fine or suspension, pending the outcome of an NBA review.
The Human Rights Campaign — the nation's largest lesbian, gay, bisexual and transgender rights group — criticized Bryant sharply.
"What a disgrace for Kobe Bryant to use such horribly offensive and distasteful language, especially when millions of people are watching," the HRC said in a statement.
"Hopefully Mr. Bryant will recognize that as a person with such fame and influence, the use of such language not only offends millions of LGBT people around the world, but also perpetuates a culture of discrimination and hate that all of us, most notably Mr. Bryant, should be working to eradicate."