Henderson was the Jacksonville Jaguars' first-round pick (9th overall) in the 2002 NFL Draft. He finished his rookie season with 53 tackles (44 solo), 6.5 sacks, 6 pass deflections, and 1 forced fumble in 16 games (13 starts). In 2003, he had 56 tackles (45 solo and 13 for loss), 3.5 sacks, 7 pass deflections, 3 forced fumbles, and 33 quarterback pressures in 16 starts. In 2004, he finished with 75 tackles (62 solo and 5 for loss), 5.5 sacks, 4 pass deflections, 1 fumble recovery, and 39 QB pressures in 16 starts and was selected to his first Pro Bowl. In 2005, Henderson had 70 tackles (52 solo and 4 for loss), 3 sacks, 7 pass deflections, 3 forced fumbles, and 20 QB pressures in 16 games (15 starts). He also started in the Jaguars' lone playoff game and recorded 5 tackles and 1 sack. On March 21, 2006, Henderson signed a new 6 year, $34 million contract extension with the Jaguars through 2011. The contract also contains $13.4 million guaranteed. Henderson responded with 51 tackles (38 solo and 6 for loss), 3.5 sacks, 6 pass deflections, and 7 QB pressures, as well as his 2nd Pro Bowl selection. He was also named an AP 2nd Team All-Pro. He was filmed by the NFL Network getting pumped up for a game by having a team trainer slap him in the face. In 2007, Henderson finished with 38 tackles (28 solo), 2 sacks, 3 pass deflections, 1 forced fumble, and 13 QB pressures in 15 starts, marking the first time in his career he did not play all 16 games. Henderson started both playoff games for the Jaguars, finishing with 9 tackles and 2 sacks. In March 2008, Henderson's good friend and fellow defensive tackle Marcus Stroud was traded to the Buffalo Bills. Henderson finished the 2008 season with 44 tackles (34 solo), 2 sacks, and 2 pass deflections in 14 games. In the 2009 offseason, Jaguars coach Jack Del Rio was critical of Henderson for pulling out of OTAs, claiming he was faking a shoulder injury to avoid having to participate. Although many believed Henderson would be released, he met privately with Del Rio and claimed that he had "bought in". Henderson finished the 2009 season with 36 tackles (26 solo), 3 sacks, 3 pass deflections, and 2 forced fumbles.
Henderson was released by the Jaguars on April 26, 2010.
On June 11, 2010, Henderson signed a one-year contract with the Oakland Raiders. On February 24, 2011, he signed a two-year, $8 million contract with the Raiders. Following the 2011 season, he was released on March 14, 2012.
Henderson is married to Aleviar Henderson and has two sons and two daughters.
In 2007, Henderson opened a clothing store in Jacksonville called "4 Big Men by Big Hen" that sold big and tall clothing. However, the store was later closed.
In 2013, Henderson opened a restaurant in Jacksonville called "Big John's Crumpy's Wings & Things". The menu features whole wings, burgers and a large selection of southern fare. However, the restaurant closed in 2015. The franchise hails from Memphis, Tennessee.