Frye was born to John H. and Helen Mushat Frye in Birmingham. During the 1930s he taught in a private school in Birmingham. He went to Princeton University to earn his B.A. (1943), M.A. (1950), and Ph.D. (1952). In 1943 he interrupted his studies to receive an officer's commission to the United States Army. He earned a Bronze Star for his service with the Army's field artillery during the Battle of the Bulge.
In 1952, Frye accepted a teaching position at Emory University in Atlanta. During his time there he worked with local officials to integrate public schools in the Atlanta area. He worked under a Guggenheim Fellowship from 1956-1957. In 1961 he joined the Folger Shakespeare Library in Washington D.C. as a researcher in residence.
He returned to teaching in 1965 at the University of Pennsylvania, remaining there until his retirement in 1983. In 1978, he co-founded the Center of Theological Inquiry, an independent institution sponsored by the Princeton Theological Seminary and also accepted the Felix E. Schelling Professorship of English Literature.
Frye was named curator of the Furness Memorial Library at Penn in 1980. He was awarded the Thomas Jefferson Medal by the American Philosophical Society in 1997. He was also an active elder at Bryn Mawr Presbyterian Church. He died at the age of 83 of complications from heart disease. At his death he was survived by his wife, Jean Elbert Steiner Frye, a son, Roland M. Frye, Jr, and a grandson, Joseph Frye.
- Frye, Roland (1955) The Teachings of Classical Puritanism on Conjugal Love.
- Frye, Roland (1960) God, Man, and Satan: Patterns of Christian Thought and Life in "Paradise Lost," "Pilgrim's Progress," and the Great Theologians. Kennikat Press.
- Frye, Roland (1961) Perspective on Man - Literature and the Christian Tradition. Westminster Press.
- Frye, Roland (1963) Shakespeare and Christian Doctrine. Princeton University Press.
- Frye, Roland (1965) The Bible: Selections from the King James Version for Study as Literature. Houghton Mifflin.
- Frye, Roland (1965) The Reader's Bible, A Narrative. Princeton University Press.
- Frye, Roland (1967) Shakespeare's Life and Times: A Pictorial Record. Princeton University Press.
- Frye, Roland (1978) Milton's Imagery and the Visual Arts: Iconographic Tradition in the Epic Poems. Princeton University Press. ISBN 0691063494
- Frye, Roland (1982) Shakespeare: The Art of the Dramatist. Routledge. ISBN 0415352894
- Frye, Roland, ed. (1983) Is God a Creationist?: The Religious Case Against Creation-Science. Scribner. ISBN 0023395605
- Frye, Roland (1984) Renaissance Hamlet: Issues and Responses in 1600. Princeton University Press. ISBN 0691065799
- Frye, Roland (1988) Language for God and Feminist Language: Problems and Principles.
- Pelikam, Jaroslav J. (June 2006) "Roland Mushat Frye". Proceedings of the American Philosophical Society. Vol. 150, No. 2