The largest party in Brazil’s governing coalition, the PMDB, has voted to pull out.
The centrist party called for an “immediate exit” from President Dilma Rousseff’s government.
The move could hasten impeachment proceedings against President Rousseff, correspondents say.
Opposition lawmakers want to remove Ms Rousseff over claims that she manipulated accounts to hide a growing deficit.
The decision comes a day after tourism minister Henrique Eduardo Alves from the PMDB – the Brazilian Democratic Movement Party – stood down.
At a leadership meeting on Tuesday, the PMDB decided that its six remaining ministers in President Rousseff’s cabinet must resign or face ethics proceedings.
The BBC’s Laura Bicker in Sao Paulo says President Rousseff could now be temporarily suspended from office by Congress as early as May.
She would be replaced by Vice President Michel Temer, leader of the PMDB, while the Senate decides if she should permanently leave her post, our correspondent adds.
Speaking before the vote, Osmar Terra, a lawmaker from the PMDB, said he had little doubt which way it would go.
“It will be a goodbye to the government,” he said.
Analysts say a considerable number of lawmakers from the PMDB have felt uneasy about their alliance with the left-wing Workers’ Party for a while.
Their unease has been compounded by calls for Ms Rousseff’s impeachment and a widening corruption scandal involving senior members in the Workers’ Party.
The president needs one third of the members of the lower house of Congress to vote against her impeachment for the proceedings to be shelved.
Without the PMDB, she could lose as many as 69 votes at once in the 513-member Chamber of Deputies.
Another of Ms Rousseff’s coalition partners, the centre-left Social Democratic Party (PSD) said it had given its lawmakers a free vote in any possible impeachment proceedings.
Lula under pressure
On Monday, Ms Rousseff’s mentor and predecessor in office, Luiz Inacio Lula da Silva, likened the moves to impeach her to a coup.
But Lula himself is under pressure.
The Supreme Court suspended his appointment as Ms Rousseff’s chief of staff earlier this month and is due to take a final decision on the matter next week.
Opponents of the government said the president had given Lula the post so he could escape investigation and possible proceedings over allegations of money laundering.
Under Brazilian law, cabinet members can only be investigated by the Supreme Court, not by lower courts.
There have been mass protests demanding the impeachment of President Rousseff in cities like Sao Paulo and Rio de Janeiro.
But there have also been rallies in support of the government by those who say the president is the victim of a campaign to drive the Workers’ Party from government.