A staple of the world championship since 1965, Automotodrom Brno last featured on the 2020 calendar.
Since 1987, the grand prix has been held on the purpose-built 5.4km circuit.
Brno hosted MotoGP annually from 1987 to 2020 with the exception of 1992.
Next year, Brno will return to the calendar on a four-year deal through to 2029.
Where Brno slots into the calendar is yet to be confirmed. The race was traditionally held in August, a date that was replaced by the Britsh Motorcycle Grand Prix at Silverstone.
Silverstone has since been moved to May, which could conceivably open up an August return for Brno.
“We’re delighted to be able to announce our return to Brno,” said Carmelo Ezpeleta, CEO of MotoGP rights holder Dorna Sports.
“It’s a classic, with a fantastic layout that has staged some really memorable moments in its long history in MotoGP.
“We’re really excited to return and stage more, with the Tissot Sprint every weekend and even more for fans to enjoy on site to get closer to their heroes than ever.
“We’ve seen the hillsides here packed with passion for our sport so many times – when it was the best-attended grand prix of the year on a number of occasions – and we’re very happy to bring MotoGP back to our fans in Czechia.”
Petr Fiala, the prime minister of the Czech Republic, hailed the return of MotoGP a major coup for the country.
“Motorcycle racing has a tradition at the Brno Masaryk Circuit dating back to the 1950s,” said Fiala.
“MotoGP is a prestigious global event that brings tens of thousands of visitors to the Czech Republic, particularly to South Moravia, many of whom come from abroad.
“I am very pleased that we have managed to continue this tradition and that the MotoGP race will return to Brno after five years.”
MotoGP has offered some insight into next year’s calendar with confirmation that the Thailand Motorcycle Grand Prix will open the season on the first weekend of March.