Johnson led 156 of the 334 laps while hoping to give owner Rick Hendrick his 200th career victory.
But he couldn’t stay with Biffle, even scraping hard into the wall trying to catch up after Biffle drove under him in Turn 3 and completed the pass before the start-finish line.
Biffle, the season points leader, went on to win the fastest Cup race at the 1½-mile, high-banked Texas track.
His average speed of 160.577 mph put his Ford 3.2 seconds ahead of the Chevrolet driven by Johnson.
It was Roush’s ninth win in 23 Cup races at Texas, and completed a Lone Star State weekend sweep. Ricky Stenhouse Jr. won the Nationwide race on Friday night for Roush’s fourth Texas victory in a row and ninth overall in the second-tier series.
Biffle got his 17th career victory. It was his first since an October 2010 race in Kansas, where the series goes next week.
After starting third, Biffle was among the lead pack the entire race, leading 91 laps on a fast-paced and windy night.
There were only two cautions for 10 slowed-down laps, both for debris, and the race finished with a record 234 consecutive laps of green-flag racing.
Mark Martin finished third in a Michael Waltrip-owned Toyota, followed by Jeff Gordon and Roush driver Matt Kenseth. Polesitter Martin Truex Jr., another Waltrip car, finished sixth after leading 68 laps.
Biffle got his eighth consecutive top-10 finish at Texas, where he had a victory in 2005 even before that stretch.
For Hendrick Motorsports, it was the second race in a row that Hendrick was close to getting his 200th victory.
Johnson, Gordon and teammate Dale Earnhardt Jr. were running in the top three at Martinsville two weeks ago before a late restart in the last race before the Easter weekend break.
But Clint Bowyer’s aggressive inside move took out Johnson and Gordon, and Earnhardt ended up third.