The party of Ukrainian President Viktor Yushchenko has said it will pull its ministers from the governing coalition, amid a rift with the prime minister.
The two leaders have been bitter rivals
Mr Yushchenko's Our Ukraine bloc said it would join the opposition.
Our Ukraine has three ministers in Prime Minister Viktor Yanukovych's government, even though it is not formally part of the coalition.
The move comes despite a pact signed between Mr Yushchenko and Mr Yanukovych only a few months ago.
Relations between the two have been deteriorating, with the cabinet refusing to endorse presidential decrees, and the president rebuking the prime minister for contradicting his stance on foreign policy.
Attempts to resolve their differences by negotiation were "finished", said Roman Bezsmertny, head of the Our Ukraine bloc in parliament.
"The Our Ukraine party is going into opposition to the government and will propose to the president that he recall ministers who are members of this party," said party spokeswoman Tetiana Mokridi.
Mr Yushchenko and Mr Yanukovych were bitter rivals during the 2004 presidential campaign, after which the former was swept to power by the Orange Revolution.
But Mr Yanukovych bounced back in parliamentary elections this year, forcing Mr Yushchenko to appoint him prime minister.
The two agreed a deal, which included having Our Ukraine ministers in the cabinet, including the foreign minister.
However the governing coalition has the support in parliament to survive without the help of Our Ukraine.