Born in Rayswick in Marion County, Kentucky, during the Civil War Knott was supportive of the North. A staunch unionist, he remained in Jefferson City, Missouri after the rest of the state government fled the Union Army as it captured the state capital.

Radical Republicans subsequently seized control of the Missouri state government. Knott remained Attorney General through the fall of 1861 when, despite his unionist sympathies, the Radicals demanded he sign a personal oath to Abraham Lincoln. Knott refused, arguing that his loyalty was to the United States Constitution and stating the oath was too stringent. In response the Radical Republicans vacated his office and imprisoned him.

After the war, Knott relocated to Kentucky where he was elected to the United States House of Representatives from 1867-1871. On January 27, 1871 he delivered a satirical speech in the House known as Duluth! or The Untold Delights of Duluth in opposition to the St. Croix and Superior Land Grant bill; the speech lampooned Western boosterism by portraying Duluth, Minnesota in fantastical and glowing language. The speech is sometimes reprinted in collections of humorous speechs or folklore, and is regarded as a minor classic. A rail car sorting yard outside of Duluth was named after Proctor Knott in 1890, and the town of "Proctorknott", now Proctor, Minnesota, took its name from those yards and from Proctor Knott himself.

Knott was elected to the House again from 1875-1883. In 1883 he successfully ran for Governor of Kentucky and held that office until 1887. He died in 1911 in Lebanon, Kentucky, and he is buried at the Ryder Cemetery in Lebanon, Kentucky. Knott County, Kentucky is named in his honor.