He headed Xander Schauffele, Shubhankar Sharma and Chris Paisley by a stroke on a day when defending champion Dustin Johnson hit a 69 that included a near ace at the 17th, his ball landing beyond the pin and spinning back almost into the hole.
In just his second PGA Tour start of the year, Oosthuizen ran off five birdies before landing a four-foot eagle at the par-five 15th, which plays more as a long par-four in the rarefied air more than 7,300 feet (2,225m) above sea level.
"I'm very confident with my driver at the moment. I've got a nice little cut going, and it's nice to be able to just aim left and swing away," Oosthuizen told Golf Channel.
"I've been swinging it pretty good the last two or three weeks."
He said he had misjudged the distances of some of his iron shots in the thin air, but not enough to cost him a bogey.
"You can get one with a short iron and it just blows your mind how far it goes and then the next hole you come up short. It's all about the ball flight you put on it."
A runaway seven-shot winner at St. Andrews in 2010, and a playoff loser in two other majors, the laid-back Oosthuizen is rarely happier than when driving his tractor on the family farm in the Western Cape Province of South Africa.
Golf takes precedence at this time of the year, though, as the sport's elite hone their games for the first major of the year at the Masters next month, while also trying to pick up a World Golf Championships title along the way.
If Oosthuizen's lead was no surprise, Paisley's lofty position perhaps was unexpected.
"It's the biggest tournament I've played in by a long stretch," the Briton said.
"To shoot a great score against the world's best players, I'm delighted. I didn't want to come here and just be happy to be part of it and make up the numbers.
"I'm surprised I shot six-under. That's surpassed my expectations but I was quietly confident I could have a good day."
Paisley won the South African Open in January - his first European Tour victory - and followed up with consecutive top-five finishes against strong international fields in the Middle East.
(Reporting by Andrew Both in Cary, North Carolina, editing by Nick Mulvenney)
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