The series' move to the cable network last year came at a time when Williams' contract was up for negotiation, and it was uncertain whether he would continue as the show's musical host.
ESPN — which begins an eight-year deal next season for Monday night rights — said in a statement that Williams will continue singing the anthem based on his hit "All My Rowdy Friends Are Coming Over Tonight."
Williams will perform with a band that includes guitarists Rick Nielsen from Cheap Trick and Steven Van Zandt from the E Street Band; keyboardists Little Richard and Bernie Worrell from Parliament Funkadelic; fiddler Charlie Daniels; and saxophonist Clarence Clemons of the E Street Band.
"When Hank says 'Are you ready for some football?' it injects a surge of energy and excitement before the Monday Night Football kickoff, and that will definitely be true on ESPN this fall," said Norby Williamson, an ESPN executive vice president for studio and remote production.
"Hank and this legendary group of musicians will give ESPN's MNF a new, high-energy opening every week of the season that will have fans in a frenzy just as the teams hit the field."
The "Monday Night Football" opening video will feature team-specific lyrics and visuals each week based on the 17 regular season ESPN matchups.
Williams has had a string of No. 1 hits including "Family Tradition" and "All My Rowdy Friends Are Coming Over Tonight."
His father, a star in the late 1940s to early 1950s, had hits like "Your Cheatin' Heart" and "I'm So Lonesome I Could Cry." He died in 1953.