Envoy Robert Pollard was called to the foreign ministry on Wednesday and handed a letter in which Malaysia said it was "offended that it has been unfairly and deliberately targeted by President Bush".
The row follows a major speech on nuclear proliferation by Bush last week in which he referred to the seizure of centrifuge parts made in Malaysia aboard a ship destined for Libya last October.
Centrifuges can be used to enrich uranium for nuclear weapons.
A Malaysian company owned by Prime Minister Abd Allah Ahmad Badawi's son Kamaluddin has admitted manufacturing the seized parts but says it thought they were for use in the oil and gas industry and did not know they were headed for Libya.
"We take exception to Malaysia being deliberately singled out in the speech, when President Bush has also clearly stated that other necessary parts were purchased through network operatives based in Europe, the Middle East and Africa," the protest statement says. "Yet he failed to name the countries hosting them.
"We are disappointed that the speech appears to question the commitment of the government of Malaysia on the issue of non-proliferation."
Asked to comment on the protest, US embassy spokesman Frank Whitaker told reporters:
"The charge had a very useful discussion with the ministry of foreign affairs and I can tell you that we are encouraging Malaysia to take the necessary steps to bring its export control in line with international standards. We believe this would prevent future proliferation activities."
Malaysia is a signatory to the Nuclear Non-proliferation Treaty.
The son of Malaysian PM Abd Allah
Badawi (C) is linked to allegation
On Wednesday, Malaysia welcomed a statement by a senior US official that Bush had not meant to implicate the government in the nuclear black market scandal.
US Undersecretary of State John Bolton said there was "certainly no whiff of an allegation in the president's statement that the government of Malaysia had the slightest thing to do with it".
He also appeared prepared to accept the explanation of the company involved, Scomi Precision Engineering (SCOPE), saying "perfectly reputable companies" could manufacture "these devices and not have any idea what they're ultimately being bound for".
Bush had linked the company with the nuclear black market run by Pakistan's scientist Abd Al Qadir Khan, who has admitted selling nuclear secrets.
Attention has also focused on a Sri Lankan businessman living in Malaysia, B.S.A. Tahir, who ordered the parts from SCOPE and was named by Bush as Khan's "chief financial officer and money launderer".
"We take exception to Malaysia being deliberately singled out in the speech, when President Bush has also clearly stated that other necessary parts were purchased through network operatives based in Europe, the Middle East and Africa"
Protest statement by Malaysian government to US embassy official
Tahir has been questioned by Malaysian police but not arrested, with the government saying it remained to be established whether he had broken any laws.
There has been no call from Washington, at least publicly, for Tahir's arrest.
The New York Times reported on Wednesday that Tahir had been a director of an investment holding company called Kaspadu, until recently owned by his wife in partnership with the prime minister's son, and had other business interests in the country.