John McKnight was born in Carlisle, Pennsylvania on October 1, 1754.  He prepared for college at the Latin Grammar School in Carlisle and went on to study at the College of New Jersey (Princeton).  He graduated in 1773 and went on to study theology under Reverend Robert Cooper.  He was licensed to preach by the Presbytery of Donegal between 1774 and 1775 and later was ordained in 1776 or early 1777, although the exact dates are not certain.  In 1776 McKnight married Susan Brown of Franklin County, Pennsylvania, with whom he had ten children.  In 1783 he was named a charter trustee of Dickinson College in Carlisle, a position from which he would resign in 1794 before moving to New York.  After moving, he became both a trustee and a professor of moral philosophy and logic at Columbia University.  Also achieving prominence within the Presbyterian Church, McKnight would be selected to preside at the Presbyterian General Assembly in 1795, which met in Carlisle.

On November 1, 1815, John McKnight was elected as the fourth president of Dickinson College.  Upon returning to his hometown, he was faced with a large debt at the college, which was still making payments to the family of Charles Nisbet due to arrears in his salary at the time of his death more than a decade earlier.  Then in December 1815,  John T. Corbin of the class of 1816 was killed in a duel with Dabney Carr Terrell, who then fled Carlisle.  The event shocked both the College and the town of Carlisle, greatly damaging the already dwindling reputation of the institution.  In light of these difficulties, the Board of Trustees decided on September 27, 1816 to temporarily suspend the business of the College.  This act ended the brief term of John McKnight as the institution's president.  Following his position at Dickinson, McKnight went on to preach in the Rocky Spring Church, serving that congregation until his death on October 21, 1823.