Luisa Ortega, a supporter turned critic of President Nicolás Maduro, has rejected her dismissal.
She will face trial for “serious misconduct”, the Supreme Court says.
Meanwhile, one of the opposition leaders, Leopoldo Lopez, has been returned from jail to house arrest.
Leopoldo López was taken from his home on August 1 and spent four days in a military jail.
South American regional bloc Mercosur has suspended Venezuela “indefinitely”, having previously placed it under a temporary ban. It says the country will not be re-admitted until the constituent assembly is scrapped and all political prisoners are released.
Another opponent of the government, Antonio Ledezma, is also back under house arrest after three days in jail last week.
Antinio Ledezma and Leopoldo Lopez had encouraged protests against the constituent assembly, which is dominated by government supporters.
In its first session on August 5, the assembly unanimously voted to remove Luisa Ortega from her post.
She was prevented from entering her office in Caracas by dozens of National Guard officers in riot gear, and left on a motorbike amid chaotic scenes.
In a statement released by the public prosecutor’s office, Luisa Ortega said President Maduro’s government was leading a “coup against the constitution”.
“I do not recognize the decision,” she said of her dismissal.
“[This is] just a tiny example of what’s coming for everyone that dares to oppose this totalitarian form of ruling.”
Venezuela’s Supreme Court, loyal to Nicolas Maduro, did not give details of the accusations against Luisa Ortega.
Luisa Ortega, who broke ranks with the government in March, had opposed the assembly’s inauguration on August 4, citing allegations of voting fraud.
Tarek William Saab, a supporter of President Nicolas Maduro, has been sworn in as her replacement.
Luisa Ortega’s removal was widely expected but the fact that it was decided on the first working day of the assembly suggested that the new body could take aggressive measures against President Maduro’s critics, correspondents say.
Mexico, Peru and Colombia have condemned Luisa Ortega’s dismissal.
President Maduro says the constituent assembly is needed to bring peace after months of protests sparked by severe economic hardship.
However, the opposition says it is a way for the president to cling to power.
The constituent assembly has the ability to rewrite the constitution, and could override the opposition-controlled parliament, the National Assembly.
Julio Borges, speaker of the National Assembly, said Luisa Ortega’s removal was illegal and showed that the country’s institutions had been “taken hostage by only one hand, only one political party” through “an undemocratic mechanism that is utterly dictatorial.”
Despite being oil-rich, Venezuela is experiencing severe shortages of food and medicines, as well as inflation in excess of 700%. Violent demonstrations since April have left more than 100 people dead.
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