He was briefly, in 1591–1592, vicar of Trumpington, a college living just outside Cambridge. In 1592, Sir Thomas Heneage, on behalf of Elizabeth I, created him vicar of Epping, Essex. In October 1595 he was appointed to the Crown living of Henton by Elizabeth, and in December 1595 Overall was appointed Regius Professor of Divinity at Cambridge. His election may have been a snub for Archbishop John Whitgift, who had adopted the Calvinistic Lambeth Articles. Overall, with Lancelot Andrewes, Samuel Harsnett, and others, had rejected these articles in support of Peter Baro, the Lady Margaret's Professor of Divinity, when on 12 January 1596 he attacked them from the pulpit. This opposition cost Baro his chair, as he failed to be re-elected in 1596. John Overall was also a friend to the erratic mystic William Alabaster (1568–1640), even throughout his years of imprisonment, and was the tutor to Robert Devereux, 2nd Earl of Essex at Trinity College. Perhaps Overall brought these two acquaintances together. Essex became Alabaster’s patron. In "Alabaster’s Conversion" we read: