Kevin McCarthy has been toppled in a right-wing revolt - the first time ever that a US House of Representatives Speaker has lost a no-confidence vote.
The final tally was 216-210 to remove the congressman as leader of the Republican majority in the lower chamber of Congress, reports BBC.
Hardliners in his party voted against him after he struck a deal with Senate Democrats to fund government agencies.
There is no clear successor to oversee the House Republican majority.
Congress has just over 40 days to agree on a deal to avoid another potential government shutdown.
Florida Republican Matt Gaetz, a Trump ally, filed a rarely used procedural tool known as a motion to vacate on Monday night to oust Mr McCarthy.
He accused the Speaker of making a secret deal with the White House to continue funding for Ukraine, amid negotiations to avert a partial government shutdown at the weekend. Mr McCarthy denies it.
At a private meeting of Republican politicians on Tuesday evening after losing his job, Mr McCarthy told colleagues he did not plan to run for Speaker again.
He later took aim at his political nemesis, Mr Gaetz, accusing him of attention-seeking.
"You know it was personal," Mr McCarthy told a news conference, "it had nothing to do with spending."
He said fundraising emails sent by Mr Gaetz amid the infighting were "not becoming of a member of Congress".
The hardliners who ousted him "are not conservatives", Mr McCarthy added.
He only became Speaker in January after a gruelling 15 rounds of voting in the chamber as Mr Gaetz and other right-wingers refused to support him.
To win over those hardliners Mr McCarthy agreed to make it possible for a single member to put forward a motion to oust him, which is exactly what Mr Gaetz ultimately did.
Mr McCarthy was supported by 210 Republicans but eight voted against him in Tuesday`s vote, joining all Democrat members.
One vote against Mr McCarthy that surprised many came from a moderate Republican from South Carolina, Nancy Mace.
She said afterwards: "I am looking for a Speaker who will tell the truth to the American people, who will be honest and trustworthy with Congress, with both parties."
Democratic House Leader Hakeem Jeffries had said in a letter to colleagues that he would not provide the votes needed to rescue Mr McCarthy.
Congresswoman Pramila Jayapal, a left-wing Democrat from the north-western US state of Washington, said before the vote: "Let them wallow in their pigsty of incompetence."
IMAGE SOURCE,GETTY IMAGES Democratic House minority leader Hakeem Jeffries refused to rescue Kevin McCarthy
The packed chamber - which Republicans control by a 221-212 majority - was mostly silent as members awaited the result of the roll call vote.
"The office of Speaker of the House is hereby declared vacant," said Arkansas Republican Steve Womack with a bang of his gavel, to audible gasps.
Earlier in the day the former president, Donald Trump, said on social media that the party should be "fighting the Radical Left Democrats" instead of each other.
North Carolina Republican Patrick McHenry, who supported Mr McCarthy, is now the interim Speaker. He put the House in recess for a week.
It is unclear if he will have the full powers of the office, or merely administrative powers and the ability to supervise a new election.
The rules do not state how long a person could fill in as an interim, though a vote on a new Speaker is planned for 11 October.
Louisiana Republican Steve Scalise and Minnesota Republican Tom Emmer have been mentioned as potential replacements for Mr McCarthy. Neither has expressed any interest in the role.
IMAGE SOURCE,GETTY IMAGESRepresentative Nancy Mace before voting Kevin McCarthy out as Speaker
White House press secretary Karine Jean-Pierre said in a statement that President Joe Biden is hoping the House would quickly elect a new Speaker, and noted that the "challenges facing our nation will not wait".
The last two Republican Speakers - Paul Ryan and John Boehner - left Congress after repeated tangles with their more conservative colleagues.
The so-called motion to vacate had only previously been used twice in the past century to remove a Speaker - in 2015 and 2010 - though never successfully.
