The court ordered a retrial for Nicholas Slatten, saying he should not have been tried alongside three others.
It also ruled that his convicted former colleagues Paul Slough, Dustin Heard and Evan Liberty be resentenced. They were all jailed for 30 years.
The killings in Baghdad's Nisoor Square sparked international outrage.
All four men were convicted in 2014 and sentenced the following year.
Slatten received a life sentence for murder while the other three were jailed for multiple counts of manslaughter, attempted manslaughter and using firearms while committing a felony.
"The court concludes that the district court abused its discretion in denying Slatten's motion to sever his trial from that of his co-defendants and therefore vacates his conviction and remands for a new trial," the US Court of Appeals for the District of Columbia Circuit said on Friday.
The ruling said that at a retrial Slatten would be able to introduce evidence alleging he did not fire the first shots.
Separately, the court said Slough, Heard and Liberty should be re-sentenced because their 30-year prison terms were too long. It overturned one of Liberty's convictions for attempted manslaughter.
The private contractors had opened fire to clear the way for a US convoy. Fourteen civilians died and another 17 were injured.
The men claimed they were under fire from insurgents.
Dozens of victims and witnesses flew from Iraq to the US for the trial.
A US appeals court has overturned a murder conviction for a former Blackwater security guard over the killing of 14 Iraqi civilians in 2007.