Eton “works very well” without admitting girls, Jacob Rees-Mogg said after the Education Secretary suggested the school should welcome female pupils.
Commons Leader Mr Rees-Mogg, who went to Eton, spoke out after Gavin Williamson said it would be a “great step forward” if the school altered its admissions policy.
Eton College is one of the UK’s most prestigious public schools, with former pupils including Prime Minister Boris Johnson.
The school, near Windsor in Berkshire, was founded in 1440 by Henry VI and Mr Johnson is the latest in a long line of Etonians who have become prime minister.
David Cameron, Harold Macmillan and William Gladstone are among the 20 prime ministers who studied there.
Mr Williamson, who attended Raincliffe Comprehensive School in Scarborough, told BBC Radio 4’s Today programme: “I would be very much in favour of Eton taking girls.
“I think that would be a good step forward.”
But Mr Rees-Mogg told MPs: “Much though I might like my own daughter to go there, I think it works very well as it is, thank you very much.”
Downing Street said it is a matter for Eton whether it chooses to admit girls.
The Prime Minister’s official spokesman said that, while single-sex schools are an important part of the education system, the Government would support Eton if it decided to change.
“We have said consistently single-sex schools are an important of our diverse education system and it is right that parents have the opportunity to make decisions about the type of school that their children attend,” the spokesman said.
“If Eton were to become a mixed-sex school we would obviously support that decision, but it is a matter for Eton as an independent school to set their own admission criteria.”