An earlier attempt to try the four in a civilian US court was halted in 2009.
New rules for Guantanamo trials have been since introduced, including a ban on evidence obtained under torture.
However, defence lawyers still say the system lacks legitimacy, because of restricted access to their clients.
US President Barack Obama tried to shut Guantanamo at the beginning of his term. But his efforts to hold Khalid Sheikh Mohammed's trial in New York foundered in the face of political and public opposition.
A small number of victims' relatives are attending Saturday's hearing at the military complex.
'Proud' of attacks
Self-proclaimed 9/11 "mastermind" Khalid Sheikh Mohammed and the four others - Waleed bin Attash, Ramzi Binalshibh, Ali Abd al-Aziz Ali and Mustafa Ahmad al-Hawsawi - are accused of planning and executing the terror attacks of 11 September 2001, which saw hijacked planes strike New York, Washington and Shanksville, Pennsylvania and left a total of 2,976 people dead.
At Saturday's arraignment, they face charges including terrorism, hijacking, conspiracy, murder and destruction of property.
They are expected to be asked to enter a plea for the first time.
The charges can carry the death penalty.
Ahead of the hearing, Jim Harrington, the civilian lawyer for Ramzi Binalshibh, told Associated Press that although his client had previously said he was "proud" of his role in the attacks he had "no intention of pleading guilty".
"I don't think anyone is going to plead guilty," he added.
The decision to hold a military rather than a civilian trial remains controversial and follows a lengthy legal wrangle over where the five men would face justice.
Another of the defendants' lawyers, James Connell, predicted the trial would take years to complete.
Khalid Sheikh Mohammed, who is of Pakistani origin but was born in Kuwait, was captured in Pakistan in 2003 and transferred to the Guantanamo base in Cuba in 2006.
During an earlier, controversial attempt to try him before a military tribunal in 2008, he said he intended to plead guilty and would welcome martyrdom.
In 2009 the Obama administration tried to move their trial into US civilian courts, but reversed its decision in 2011 after widespread opposition.
The five were eventually charged in June 2011 with offences similar to those they were accused of by the Bush administration.
The Pentagon has previously said Khalid Sheikh Mohammed admitted he was responsible "from A to Z" for the 9/11 attacks.
US prosecutors allege that he was involved with a host of other terrorist activities.
These include the 2002 nightclub bombing in Bali, Indonesia, the 1993 World Trade Center bombing, the murder of American journalist Daniel Pearl and a failed 2001 attempt to blow up an airliner using a shoe bomb.
Khalid Sheikh Mohammed has alleged that he was repeatedly tortured during his detention in Cuba.
CIA documents confirm that he was subjected to simulated drowning, known as waterboarding, 183 times.