Gaspare Spatuzza, who admits having murdered dozens of people, will on 5 October give evidence at the trial of Gen. Mario Mori and Col. Mauro, who are accused of abetting Provenzano.
Provenzano was arrested at a hideout in a Sicilian farmhouse in 2006 after 40 years on the run.
Prosecutors at Mori's trial will next month question Spatuzza about the Sicilian mafia's 1992 bombing campaign which killed leading anti-Mafia prosecutors Giovanni Falcone and Paolo Borsellino in Palermo and 22 people in Florence and Rome.
Prosecutors will also quiz Spatuzza on a failed 1993 attack that targeted paramilitary Carabinieri police at Rome's Olympic Stadium.
Spatuzza was in 2011 granted police protection after he testified in 2009 that jailed Sicilian mafia boss Giuseppe Graviano told him in 1994 that then-prime minister and media mogul Silvio Berlusconi was helping the mafia.
In his testimony, Spatuzza also made accusations about conservative senator Marcello Dell'Utri, a close Berlusconi associate, saying the mafia had "obtained everything" from him and from Berlusconi. He and Dell'Utri vehemently deny Spatuzza's allegations.
Dell'Utri is currently at the centre of a probe into allegations he extorted 40 million euros from Berlusconi and is set to have a retrial after receiving a seven-year jail sentence for abetting the mafia.
Italy's top appeals court, the Court of Cassation in March ordered the retrial, saying he had not received a fair trial.
But in its explanation of the decision, the court said there was evidence that Dell'Utri had acted as intermediary with the mafia for Berlusconi, handing it "substantial sums" of so-called protection money.