Becoming the first team since the New England Patriots in 2004 to win successive Super Bowl crowns, Andy Reid's team dramatically forced overtime with three seconds of the fourth quarter remaining, only for Jake Moody's field goal to ostensibly carry the day for the 49ers.
However, as the clock ticked down into the final few seconds of the overtime period, the irrepressible Patrick Mahomes picked out Mecole Hardman with a three-yard touchdown pass to spark pandemonium for those of a Chiefs persuasion.
After going 48 years without a single Super Bowl next to their name from 1970 to 2018, the Chiefs have now come up trumps in three of the past five NFL seasons, although it took a monumental effort from Reid's charges to deny the 49ers their moment in the spotlight.
Defences came out on top in a scoreless first quarter, before Moody's opening field goal and a 21-yard pass from Jauan Jennings to Christian McCaffrey propelled the 49ers into double figures before
Harrison Butker cut the NFC champions' lead down to seven.
Frustrations were boiling over for the Chiefs, as Travis Kelce was taken off and gave coach Reid an earful after a fumble from Isiah Pacheco, causing the 65-year-old to momentarily lose his balance before Kelce was ushered away.
Nevertheless, the reigning champions kicked into gear in the third quarter and led for the first time in the match thanks to Mahomes's 16-yard pass to Marquez Valdes-Scantling in the end zone, giving Reid's men a three-point advantage heading into the final quarter.
However, after throwing the game's first touchdown pass for McCaffrey, Jennings received a 10-yard delivery from Brock Purdy to restore the 49ers' slender advantage, becoming the first player in seven years to throw and receive a touchdown pass in a Super Bowl match.
Moody's kick for an extra point was blocked, though, and while the 24-year-old did find the mark again from 53 yards, two efforts from Butker through the posts forced overtime for just the second time in the Super Bowl.
With just over seven minutes remaining, Moody propelled the 49ers back into a three-point lead from 27 yards, but on a first down in the final 10 seconds, Mahomes arrowed a straightforward three-yard delivery to Hardman to break 49ers hearts.
Chiefs quarterback Mahomes was bestowed with the Super Bowl's Most Valuable Player award in Las Vegas, becoming the first player across the NFL, NBA, MLB and NHL to take three championship MVP awards in his first seven seasons.