He was born in Norwich, Connecticut, the son of James Dwight and his wife Susan Breed, and the grandson of Yale President Timothy Dwight IV.
He received his undergratuate degree at Yale in 1849.
Appointed professor of sacred literature at Yale, he assisted in the reorganization of the divinity school, edited the New Englander (1866–74), and served on the American committee on the revision of the Bible (1873–85). In 1886 he succeeded Noah Porter as president of Yale. He expanded the institution, securing the legislative charter that authorized the title university instead of college, and retired in 1898.
He is the author of Thoughts of and for the Inner Life (1899) and Memories of Yale Life and Men (1903).