U.S. District Judge Henry E. Hudson is the first federal judge to strike down the law, which has been upheld by two other federal judges in Virginia and Michigan. Several other lawsuits have been dismissed and others are pending, including one filed by 20 other states in Florida.
The government had argued the Commerce Clause of the Constitution gives the government the power to require individuals to buy health insurance or face a penalty, a provision due to take effect in 2014.
But Hudson sided with Virginia Attorney General Kenneth Cuccinelli in saying the mandate overstepped the Constitution.
"This case, however, turns on atypical and uncharted applications of constitutional law interwoven with subtle political undercurrents," Hudson wrote. "The outcome of this case has significant public policy implications. And the final word will undoubtedly reside with a higher court."
There was no immediate comment from the White House.
The Department of Justice, which defended the law in court, stood by its argument that Congress was within its rights to enact the law.
"We are disappointed in today's ruling but continue to believe - as other federal courts in Virginia and Michigan have found - that the Affordable Care Act is constitutional," said Justice Department spokeswoman Tracy Schmaler.
The lawsuit was filed by Cuccinelli, a Republican, in defense of a new state law that prohibits the government from forcing state residents to buy health insurance.
Cuccinelli argued that while the government can regulate economic activity that substantially affects interstate commerce, the decision not to buy insurance amounts to economic inactivity that is beyond the government's reach.
"This won't be the final round, as this will ultimately be decided by the Supreme Court, but today is a critical milestone in the protection of the Constitution," Cuccinelli said in a statement after the ruling.
Hudson, a Republican appointed by President George W. Bush, sounded sympathetic to the state's case when he heard oral arguments in October, and the White House expected to lose this round.
Administration officials told reporters last week that a negative ruling would have virtually no impact on the law's implementation, noting that its two major provisions - the coverage mandate and the creation of new insurance markets - don't take effect until 2014.