The billionaire couple has said they will continue to work together on the Bill and Melinda Gates Foundation - one of the world's largest private charitable organisations - despite their split.
The organisation has spent billions fighting causes such as infectious diseases and encouraging vaccinations in children.
Taking to Twitter, Bill and Melinda Gates wrote: “After a great deal of thought and a lot of work on our relationship we have made the decision to end our marriage.
"Over the last 27 years, we have raised three incredible children and built a foundation that works all over the world to enable all people to lead healthy, productive lives.
"We continue to share a belief in that mission and will continue our work together at the foundation, but we can no longer grow together as a couple in the next phase of our lives.
"We ask for space and privacy for our family as we begin to navigate this new life.”
The couple met in the late 1980s when Melinda joined Bill's Microsoft firm and they have three children together.
The duo were creators, along with Warren Buffett, of the Giving Pledge, which involves giving away more than half of their wealth.
According to Forbes, Bill Gates – co-founder and former CEO of Microsoft – is the fourth wealthiest person in the world.
The announcement from the Gates comes two years after Jeff Bezos, CEO of Amazon, and his wife, MacKenzie, announced they were getting divorced.