The Houston Oilers drafted Matthews with the ninth overall pick in the first round of the 1983 NFL Draft. In Houston, he blocked for the legendary Earl Campbell and eventually played all line positions (guard, center and tackle), going to the Pro Bowl as a guard and center. He was selected to fourteen Pro Bowls in all, tying a league record held by Merlin Olsen. Matthews was also named First-team All-Pro nine times (1988–1993, 1998–2000) and All-AFC 12 seasons (1988–1993, 1995–2000). He was selected as a guard on the NFL’s All-Decade Team of the 1990s. Matthews spent his entire career with the Oilers franchise, which relocated after the 1996 season and became the Tennessee Titans. An extremely durable player, Matthews retired after the 2001 season having played more games (296) than any NFL player, excluding kickers and punters (since surpassed by Brett Favre; Matthews still holds the record for linemen), and played in more seasons (19) than any offensive lineman. He never missed a game because of injury (the 1987 season was shortened due to a player strike), and started 229 consecutive games. In 1999, the Titans made it to Super Bowl XXXIV in which Matthews started, however they lost to the Kurt Warner-led St. Louis Rams. Matthews is the only player who played against the Baltimore Colts in their last game at Memorial Stadium and against the Baltimore Ravens in their last game at Memorial Stadium.
In his first year of eligibility, Matthews was elected to the Pro Football Hall of Fame as part of the Class of 2007. He was inducted during the Enshrinement Ceremony on August 5, 2007 with the unveiling of his bust, sculpted by Scott Myers. He is the only player from the Tennessee Titans to be given this honor since the relocation from Houston. He was the fifth player from the 1983 NFL draft class to be enshrined, joining Dan Marino, Eric Dickerson, John Elway, and Jim Kelly; Darrell Green and Richard Dent later became the sixth and seventh.
On February 27, 2009, Matthews returned to Houston where he was signed on as an Offensive Assistant with the Houston Texans after volunteer coaching at his children's high school, Elkins High School. On February 9, 2011, Matthews was hired as offensive line coach by new Tennessee Titans head coach Mike Munchak. Both were Hall of Fame lineman for the Houston Oilers. Regarding his new job, Matthews stated, "For me this is an opportunity of a lifetime. It is such a unique opportunity to work with Mike because I think he will do a great job. It is just one of those things I couldn't pass up."
After finishing with a record of 7-9 for the 2013 season, Titans general managerRuston Webster and Tommy Smith, who took over as CEO after owner Bud Adams died on October 21, 2013, met with Munchak and gave him the option to fire a large contingent of assistant coaches, which included Matthews, in exchange for an extension and a raise, or lose his job as head coach. Munchak was not willing to fire everyone they were ordering him to fire, so Munchak parted ways with the Titans (after a combined 31 years with the franchise as a player, assistant coach, and head coach), along with Matthews and the other assistant coaches they wanted him to fire.