Speaking to parliamentary deputies in Tehran on Tuesday, Ghulam Ali Haddad-Adil said it was not for France, Germany or Britain to decide what was in the best interests of Iran.
The additional protocol would allow much tougher UN inspections of nuclear facilities than are currently undertaken.
"I say to France, Germany and Britain not to tell the Iranian parliament what to do."
He was referring to a tough draft resolution put to the board of the UN's International Atomic Energy Agency by the European Union's leading three nations, currently under discussion in Vienna.
"The Iranian parliament does not take orders from foreigners, because these orders do not reflect the interests of the Iranian people. If we consider it to be in the interests of the Iranian people we will adopt it, if not we will not," he said.
The parliament speaker also warned the Europeans not to "fall into the trap of the Zionists" by pressuring the Islamic republic over its nuclear programme.
EU resolution considered
The highly critical resolution being debated by the board and drafted by the EU troika of Britain, Germany, and France states that the board "decides to remain seized of the matter".
But the EU document criticising Iran contains no deadline or trigger for sanctions if Tehran does not meet the demands laid out.
"The Iranian parliament does not take orders from foreigners, because these orders do not reflect the interests of the Iranian people"
Ghulam Ali Haddad-Adil,
Iran parliament speaker
The US is said to be calling for tougher wording and some kind of deadline.
On Monday, the UN's chief nuclear inspector, Muhammad al-Baradai, voiced his exasperation at the slow pace of the international inquiry into Iran's nuclear capabilities.
He contradicted Iran's assertions that it was cooperating fully with the International Atomic Energy Agency and declared that the jury was still out on whether Iran was building a nuclear bomb.
He also said Iran had not yet satisfactorily explained traces of enriched uranium found last year on its equipment by the inspectors.
The board meeting, expected to last at least three days, came after the submission of a 21-page report by al-Baradai in which he cited fresh evidence that Iran had embarked on an illicit uranium-enrichment project which could produce bomb-grade nuclear fuel.