Johnson went around Deere Run in a seven-under 64 to sit atop the leaderboard alongside Colombia's Camilo Villegas.
Matt Bettencourt, Daniel Summerhays and South African Brendon de Jonge were a stroke behind.
Johnson is searching for his first PGA Tour win since his success here last year and quickly returned to the form that earned him last year's title.
The 37-year-old shot a flawless round of seven birdies with his playing partners Steve Stricker, who won this event three years running before Johnson's win 12 months ago, and Davis Love III both carding four-under 67s.
Love's round ensured he remained in contention to continue his record of having played in 26 consecutive Open Championships since his debut in 1987.
The 49-year-old will not, however, play at Muirfield next week unless he wins in Illinois.
Overcoming Johnson and Villegas will pose a major hurdle though.
Villegas was impressive as he blotted his copybook only with a bogey on the par-four ninth.
The 31-year-old, without a Tour win in over three years now, recovered purposefully with birdies on four of his next five holes to shoot up the standings.
Johnson's round continued his fondness with a course that he has now shot 72 under on in total since 2009.
American's Vaughn Taylor, Robert Streb, Jim Herman, Kevin Streelman and Boo Weekley were on five under along with Australia's Matt Jones and David Hearn of Canada.