A young man named Wesley Gibson (James McAvoy) works at a dead-end job with an overbearing boss. He takes anti-depressants when he feels stressed out. His live-in girlfriend is sleeping with his best friend. He speaks of how his father left when he was just one week old. He wonders if maybe when he was born his father looked into his eyes and saw a failure.
Elsewhere, a man called "Mr. X" (David O'Hara) meets with a ballistics expert to find out who made a particular bullet for a "competitor". She observes that the bullet is "clean", i.e., not traceable to a particular gun. (As becomes clear later, this is because it is the last stage of a multi-stage bullet.) Suddenly, a sniper shoots the ballistics expert in the head from a nearby building. Mr. X leaps through the window and shoots his opponents in mid-flight, killing them. He lands on the building and begins talking to a man on a cell phone, unaware that he is standing on a marked spot. He notices it as the man, named Cross fires a multi-stage bullet from across town, killing Mr. X by going through the back of his head and out through his forehead (the flight of the bullet is shown in slow-motion, backward).
One night at a pharmacy, Gibson meets a mysterious woman who tells him his father was an elite assassin who had been killed the day before. Gibson replies that his father abandoned him a week after his birth. At that moment, Cross appears, gun in hand. The woman opens fire on Cross. Gibson and the woman escape from the resulting shoot-out and have a wild car chase in the streets of Chicago. The woman brings Gibson to the headquarters of The Fraternity, a thousand-year-old secret society of assassins. The group's leader, Sloan (Morgan Freeman), formally introduces Gibson to Fox (Angelina Jolie), the woman from the night before, and invites him to follow in his father's footsteps as an assassin. Sloan tests Gibson by making him shoot the wings off a fly. When Gibson refuses, a gun is put to his head, triggering a panic attack. Gibson somehow manages to shoot the wings off several flies. Sloan says that he was able to do that because his heart beats 400 times a second when he's stressed. When Sloan asks him whether he want to know how to control it, he runs away in fear. Gibson wakes up the next day hoping everything was a dream, but discovers his father's gun (which he stashes in the toilet tank), and that he has $3.6 million in his bank account. At work, Gibson tells off his boss, bashes his duplicitous friend with a computer keyboard (forming the words "FUCK YOU" with the f, u, c, k, y, o letters and a tooth that pops out of his friend's mouth), and storms out. Gibson then sees pictures of himself and Fox on the front page of several newspapers as wanted fugitives for the pharmacy shooting. Then he notices Fox, who has been waiting outside, and she gives him a ride back to the Fraternity headquarters - an unassuming textile mill.
Sometime later when his training is complete, Gibson is given orders to kill people from the Loom of Fate, a loom that gives the names of the targets through a binary code hidden in weaving errors of the fabric. While on his first assignment, Gibson has second thoughts and hesitates killing his target. In a flashback we learn that he told Fox it isn't right to kill people without knowing anything about them or why they deserve to die. Fox then relates a childhood story, about a judge handling a sensitive case, and the defendant had ordered him assassinated. A hired killer held the young girl at knifepoint as they waited for her father to return home. The killer then lit the father on fire as the young girl watched, and then branded his initials into her neck. Fox explained that the man who killed the judge had been targeted by the Fraternity several weeks prior to the events of the story, but their assassin had failed to carry out his duty. Fox then tells Gibson The Fraternity's idea, "Kill one and maybe save thousands". As she prepares to leave, he notices initials branded on her neck and realizes the story was about her. Back to present we see Gibson bends a bullet trajectory to kill the target a moment after this recollection.
Whenever Sloan orders Gibson to kill a person, Gibson would ask whether the target is Cross as he cannot wait to have revenge. At one time, Sloan grants his wish as the next target is Cross. Fox feels that it is too early but Sloan gives her another order where the target is Gibson.
Gibson and Fox travel to the Fraternity's original base of operations in Europe. The two easily capture Pekwarsky and force him to take them to Cross. The meeting leads to a confrontation between Gibson and Cross on a moving train. Fox steals a car and crashes it into the train, eventually causing the train to derail when it reaches a bridge over a deep ravine (killing all innocent passengers). Gibson is about to fall into the ravine before Cross catches his hand, saving his life. Gibson unhesitatingly shoots him. Before Cross dies, he tells Gibson that he is his real father and that the Fraternity had been lying to him. Fox confirms the truth and explains that Gibson was recruited because he was the only person that Cross wouldn't kill. Fox then tells Gibson about the kill order on him and raises her weapon to shoot him. Gibson, however, shoots the glass underneath him and plunges into the river below.
Gibson awakes in an apartment across the street from his former apartment. He finds Pekwarsky there. Upon inspecting the apartment, he discovers it belonged to his father, who had been monitoring him his whole life. Pekwarsky hands Gibson a loom weaving and tells him to decode it. Gibson is shocked to discover Sloan's name in the weaving. Pekwarsky explains that after Sloan discovered that he was the next target stated by the Loom of Fate, he started manufacturing his own targets and after discovering this Cross goes rogue and Sloan turns the Fraternity against him. Since then Sloan has used false kill orders to direct the Fraternity as mere contract killers. Gibson realizes that Cross had never actually tried to kill him in their previous confrontations; he had been assassinating Fraternity members to keep them away from Gibson. Pekwarsky departs after giving Gibson plane tickets, stating that his father wished him a life free of violence. While exploring the apartment further, Gibson discovers a secret room containing all of his father's weapons and maps. He even finds a supply of the Exterminator's mini-bombs, realizing that the Exterminator had been working with his father. Gibson then devises a plan to take out Sloan and the Fraternity. Upon entering Sloan's office, he finds himself surrounded by Fox and her fellow master assassins. Gibson tells them that Sloan is killing for profit by providing his killers with fraudulent kill orders. He then attempts to kill Sloan, but is disarmed by Fox.
Fox asks Sloan if this is true. Sloan then reveals that all of their names had come up in the weaving, and that he had merely acted to protect them. He then goes on to explain that if they truly believe in the code then they should take their lives right where they stand. Otherwise, they should kill Gibson. The other assassins decide to kill Gibson, but Fox turns on her fellow assassins. She "curves" a bullet to kill the assassins who had been standing in a circle, then throws her gun to Gibson before stepping back into the bullet. Sloan escapes. Gibson, penniless once more, does not know what to do with himself. While Gibson provides a voice-over, the audience sees a young man sitting in front of a computer in a cubicle much like Gibson did at the beginning of the film. The man types the name "Wesley Gibson" into Google and searches for it but does not have any results, as in the beginning of the film. Sloan appears and points a gun at the man's head. At that moment, the man turns around and is revealed to be a decoy and looks down. Sloan also looks down and realizes he is standing on a marked spot. He then looks up and says, "Oh, fuck", before Gibson, who is actually miles away, shoots him in the head from the comfort of his own apartment, from the same window his father killed Mr. X at the beginning of the movie. It is also shown that the same bullet passed through an energy drink his former best friend was holding as his unfaithful girlfriend looks on in shock, and passed through a donut his former boss was about to eat mere moments before it killed Sloan.
The movie ends with Gibson breaking the fourth wall, addressing the audience and giving an overview of his last six weeks as an assassin saying, "This is me taking back control of my life. What the fuck have you done lately?"