Lee Ki-woo started out as a model before debuting as an actor in 2003 through director Kwak Jae-yong’s romance melodrama movie The Classic. In 2005, he appeared in the films Tale of Cinema and Sad Movie. That same year, he joined the main cast of the KBS2 series A Love to Kill, which was named Best Drama Series at the 2006 Asian Television Awards. In 2008, he starred with Choi Ji-woo and Yoo Ji-tae in SBS’s A Star’s Lover. In 2011, he played one of the main characters in the tvN romantic comedy Flower Boy Ramyun Shop. In 2016, he received the Hot Star Award at the 9th Korea Drama Awards for his appearance in the tvN melodrama Memory. In 2017, he earned an Excellence Award nomination at the 10th Korea Drama Awards for the high-rating JTBC series Woman of Dignity.

Lee Ki-woo enlisted for his mandatory military service in November 2009. He served an active duty at the Armed Forces Seoul Hospital until September 2011. He dated his Flower Boy Ramyun Shop co-star Lee Chung-ah.