Officials at the Dallas Zoo in Texas said they have no idea what caused the attack.
The pride of five -- two brothers and three sisters -- had lived together "peacefully" for years without incident, the zoo said.
Then on Sunday, one of the males wrapped his jaws around the neck of a 5-year-old lioness named Johari. She died quickly.
"Johari was a remarkable animal, as are all of our lions," said Lynn Kramer vice president of animal operations and welfare at the Dallas Zoo.
More From This Section
Visitor Michael Henshaw described the shocking scene.
"At first you think they're playing; then you realize he's killing her... And you're watching it," Henshaw told WFAA news. "You just can't believe your eyes."
Visitor Dylan Parker described the attack as eerily calm as the lion just lay beside Johari "and held her by the neck for like 10 minutes... Just holding it there, waiting until it quit moving."
Zoo officials said they may never know the reason for the attack, but noted that lions do sometimes kill members of their pride in the wild.
They plan to keep the lions apart while the attack is investigated but will "absolutely not" euthanize the killer lion.
"We are heartsick," the zoo said in one of a number of tweets sent in response to concerned messages.