Johnson’s bid to have his name etched onto the Claret Jug for a second time was enhanced by a 67 which earned him the early clubhouse lead on six under, and his score was later matched by overnight-leader and Carnoustie housemate Kisner, who added a 70 to his opening 65 but cost himself the outright lead with a double-bogey at 18.
Fleetwood fired the first bogey-free round of the tournament as his 65 in the morning rain propelled him into a share of third, just one stroke off the pace, while McIlroy also performed well in the damp early conditions and matched his first-round 69 to close on four under.
Defending champion Jordan Spieth is a further shot behind, and Tiger Woods will look to make an early move on Saturday from level par, but Dustin Johnson and Justin Thomas – the world’s top two players – headlined a star-studded list of names to crash out.
Zach Johnson was first into the clubhouse on six under after his bogey at the first would prove to be his only glitch of the day, the 2015 winner getting back on track with three birdies in four holes from the third before picking up further gains at 14 and 18.
The 42-year-old looked likely to be surpassed by Kisner when he raced to eight under after mixing five birdies with two bogeys over the first 14 holes, but he then gouged his second from the rough at 18 into the Barry Burn short of the green and the resulting six dropped him into a tie for the lead.
The fine weather of the first round gave way to dark skies and persistent rain as the first groups teed off, but the conditions did not phase Fleetwood, who staged a spirited recovery from his first-round 72.
The US Open runner-up made a confident start as he followed three pars with birdies at the fourth and fifth, and another at the ninth took him to the turn in 33 before he got further putts to drop at 11 and 14.
And the 27-year-old, who fired a course-record 63 at Carnoustie during last season’s Alfred Dunhill Links Championship, capped an excellent day with a 12-foot putt for a closing three at the last to move to five under for the tournament.
The American presence at the top end of the leaderboard was further boosted by Pat Perez (68) and Xander Schauffele (66) as they joined Fleetwood in a tie for third, while the rain forced McIlroy to abandon his aggressive approach as he ended the day two off the lead.
McIlroy parred the first six holes before converting a fine approach to eight feet at the seventh, and he picked up another at the 10th before a blocked second to the 12th cost him his first bogey of the round.
He responded with a good tee shot in close to set up birdie at 13 and he added another at the long next despite finding a fairway bunker with his drive, although he faltered again at 15 and avoided a further blemish at the 17th when he got up-and-down from just inside 100 yards.
McIlroy just missed out on a closing three as he finished on four under alongside two more Americans, Matt Kuchar and Tony Finau, and South African duo Erik Van Rooyen and Zander Lombard, with Spieth and another member of the “frat house” – Rickie Fowler – going into the weekend just three behind their housemates.
Just 28 players completed 36 holes in red numbers, but Woods was not among them as he offset four birdies with as many bogeys, including two dropped shots over the first three holes while endangering the spectators with a botched escape from the rough at the second.
Woods was joined on level par by long-time rival and 2013 Open champion Phil Mickelson, while world No 3 Justin Rose narrowly avoided adding his name to the list of high-profile casualties when he holed a superb 10-foot putt for his first birdie of the day at the last to scrape into the weekend with nothing to spare on three over.
But Dustin Johnson crashed to six over after a bogey, double-bogey finish to a 72, while Thomas plummeted from two under to four over in the space of only three holes as he carded three consecutive double bogeys in a torrid front nine.
World No 5 Jon Rahm also dropped six shots in three holes to make an early exit along with the 2007 champion and runner-up, Padraig Harrington and Sergio Garcia, while Bubba Watson and Ian Poulter will also have the weekend off.