Brustein's parents were Max, a businessman, and Blanche (Haft) Brustein. He was educated at Amherst College, where he received a B.A. in 1948, and Columbia University, where he received an M.A. in 1949 and a Ph.D. in 1957. During this time, he served in the Merchant Marine on tanker and Victory ships, and later at Kings Point Academy on Long Island. He also held a Fulbright Fellowship to study in the United Kingdom from 1953 to 1955, where he directed plays at the University of Nottingham. After teaching at Cornell University, Vassar College, and Columbia, where he became a full professor of dramatic literature in the English department, he became Dean of the Yale School of Drama in 1966, and served in that position until 1979. It was during this period that he founded the Yale Repertory Theatre.

In 1979, Brustein left Yale for Harvard University, where he founded the American Repertory Theatre (ART) and became a Professor of English. He served for twenty years as Director of the Loeb Drama Center where he founded the Institute for Advanced Theatre Training at Harvard. He retired from the Artistic Directorship of ART in 2002 and now serves as Founding Director and Creative Consultant.

As the Artistic Director of Yale Rep from 1966 to 1979, and of ART from 1980 to 2002, Brustein supervised over 200 productions, acting in eight and directing twelve.

Brustein has been the theatre critic for The New Republic since 1959, and is the author of fifteen books on theatre and society:

• 1964: The Theatre of Revolt: An Approach to Modern Drama (Little, Brown) ISBN 0929587537 – essays on Ibsen, Stringberg, Chekhov, Shaw, Brecht, Pirandello, O'Neill, and Artaud and Genet, considered a "standard critical text on modern drama"

• 1965: Seasons of Discontent: Dramatic Opinions 1959-1965 (Simon and Schuster) ISBN none – "an assemblage of his best magazine pieces from 1959 to [1965]"

• 1969: The Third Theatre (Knopf) ISBN 0671205374 – "a collection of pieces written between 1957 and 1968 ... that deal not only with theatre but also with literature, culture, and the movies" (from the Preface).

• 1971: Revolution as Theatre: Notes on the New Radical Style (Liveright) ISBN 0871402386 – examines campus turmoil, radicalism versus liberalism, the fate of the free university, the new revolutionary life style, the decadence of American society, and the sentimentality and false emotionalism of radical alternatives

• 1975: The Culture Watch: Essays on Theatre and Society, 1969-1974 (Knopf) ISBN 0394498143 – "As far as these bristling exhortations go, well, you have to wish the gadfly well"

• 1980: Critical Moments: Reflection on Theatre & Society, 1973-1979 (Random House)