In this sequel with basically the same characters as in the original sleeper hit, amnesiac CIA agent Jason Bourne (Matt Damon) found himself haunted by flashbacks and nightmares of "bits and pieces" of his former life (and a botched mission after he was trained as an assassin - his first 'hit' job ever). He was living two years later in Goa, India with German girlfriend Marie Kreutz (Franka Potente); he was recording his recollections in a journal to help him piece together his past (later discovered to be a crucial assassination carried out in a Berlin hotel room).
Meanwhile, in Berlin, Germany, there was a CIA sting operation, led by ambitious CIA Deputy Director Pamela Landy (Joan Allen). It was being conducted to obtain what were called the "Neski files." A paid Russian assassin named Kirill (Karl Urban) sabotaged the operation - he first planted two bombs on conduits leading from the basement's electrical circuit box - one that didn't explode had Jason Bourne's fingerprints on them, in order to frame him. Then, when the other bomb exploded to cut off the power, Kirill murdered the CIA case officer/agent and his Russian mole contact during the exchange of money ($3 million in cash) for the sale of files containing incriminating evidence about the identity of the agent who stole $20 million from the CIA seven years earlier ("These files will prove who stole your money").
Immediately afterwards, Kirill sold the files to shady Moscow oil tycoon, the Pekos Oil Company CEO named Yuri Gretkov (Karl Roden). [Spoiler: Yuri bought oil leases for $20 million with the stolen CIA money and was in a covert alliance with evil mastermind and veteran CIA boss Ward Abbott (Brian Cox). Both Conklin (Chris Cooper) and Abbott, Bourne's controllers, were involved in a secret criminal conspiracy which Neski had documented, netting them funds in their own personal bank accounts.]
Kirill's next paid mission was to eliminate Bourne in India, where he was sent by Gretkov. He made an assassination attempt on his life in the village market, but Marie was the one who was tragically shot by the sniper's fire and was killed as they were pursued, and their vehicle toppled off a bridge into a river. Bourne's attempts at artificial resuscitation were in vain. Kirill believed he had killed his target and was paid off. Undetected, Bourne escaped and decided to re-enter the "grid" to find out the plot against him and his role in Treadstone (from the first film), while keeping only one picture of him with Marie.
When Landy learned that the fingerprint at the scene of the double murder was linked to the CIA's top-secret Treadstone project (and agent Jason Bourne who was their number one operative, but had screwed up a job off the coast of France), she was condescendingly told at CIA headquarters at Langley, Virginia by Abbott that she "might have wandered a little bit past your pay grade." However, when she disclosed she was given clearance to all information from CIA Director Martin Marshall (Tomas Arana), she was told that Treadstone was "a kill squad, black-on-black" that was closed down two years ago, and Bourne's whereabouts were now unknown. A CIA substation in London picked up Bourne's reentry point into Europe at Naples, Italy, but shortly thereafter, he escaped from custody.
Landy continued her fact-gathering, learning that seven years earlier, $20 million of CIA funds disappeared during a wire transfer through Moscow. In the subsequent investigation, the CIA was contacted by Russian politician Vladimir Neski, claiming that the CIA had a leak and that they'd been ripped off by someone internally. A final determination was never reached, since Neski was killed (reportedly by his wife in Hotel Brecker in Berlin, determined at the time to be a murder-suicide) during negotiations for a meeting. Landy described how a second source, another Russian in Berlin, had recently obtained access to the Neski murder files - and they thought they could get more information by buying the files in Berlin, but Bourne (connected by fingerprints) intervened and allegedly committed a hit on the two.
The elusive Bourne, knowing that he had been framed and suspected of Kirill's double murders in Berlin, and knowing that he was being simultaneously pursued by the CIA (since he was an embarrassment to them - Marshall vowed: "We're gonna find this son-of-a-bitch and take him down") and Russian criminal Kirill (to finish the job), he stole a car and drove to Munich, Germany (on his way to Berlin), to attempt to clear his name. In Munich, he questioned former Treadstone agent Jarda (Marton Csokas), who admitted that after Treadstone was shut down, following Conklin's death in Paris: "We're the last two." When they fought together, Bourne strangled Jarda and blew up his apartment, allowing Bourne time to escape.
Bourne drove further onto Berlin, where he located Pamela Landy staying at the Westin Grand Hotel. He followed her to CIA headquarters, where she was leading a massive search effort for him. While sighting her in his sniper scope, he spoke to Landy on the phone, learning that Treadstone was shut down two years earlier, and that he had "killed two people" in Berlin. For a moment, he thought she was referring to Vladimir Neski and his wife's death, not the recent sting operation. He volunteered to be brought in, through a rendezvous with Nicky Parsons (Julia Stiles), Treadstone's operative in Paris two years earlier. After eluding snipers and other operatives, he was able to interrogate Nicky (who was wired), and realized he had been set up in Berlin (he was in India watching Marie die at the time). He angrily asked about the CIA's efforts to get him: "Why are you trying to frame me?" She also admitted that Abbott ran Treadstone and "Conklin reported to him," and that the Berlin sting buy had "something to do with a Russian politician [Neski]."
One of Abbott's young staff members, Danny Zorn (Gabriel Mann), confided in his boss about how he had investigated the two charges on the electrical box, and how the second one was deliberately not set to go off, to frame Bourne with his fingerprints ("It was staged"). When Zorn asked crucial questions: "What if somebody were trying to cover their tracks by blaming Conklin and Bourne? What if Bourne didn't have anything to do with this?", Zorn lost his life with a knife to the midsection when Abbott silenced him over this crucial bit of information.
Bourne had been in Berlin for a previous Treadstone job (although Nicky only knew of a job in Geneva) - he 'remembered' his "first time" job (for boss Conklin) killing a Russian couple, Vladimir Neski (a Russian government official) and his wife, in their Hotel Brecker room (#645) in Berlin, who were about to identify the mole thief - leaving Irena as their young orphaned daughter. When he revisited the hotel, Bourne flashbacked to the night of the Neski murders, remembering how he shot them and made it look like a murder-suicide. He then escaped pursuit from the Berlin police.
When Zorn's body was discovered, Abbott felt trapped, and phoned Gretkov from his Westin Grand Hotel room, telling him: "They're onto Neski...They can't prove anything without Bourne. Kill Bourne and you kill this investigation." Abbott reminded Gretkov that they had both stolen and split the money, but that his earlier order to kill Bourne was still paramount: "You bought those oil leases with $20 million in stolen CIA seed money. You owe me...The plan can still be salvaged. Just get Bourne. Do you hear me? He's still out there. Now get him."
Bourne had overheard the entire conversation in the hotel room, and was taping Abbott as he confessed and self-incriminated himself in the attempted conspiratorial extortion scheme, and his link to the Neski murders Bourne was tempted, but didn't kill Abbott, explaining that non-violent Marie wouldn't have wanted him to: "She wouldn't want me to. That's the only reason you're alive." The tape was mailed to Landy, and Abbott shortly afterwards committed suicide (after vowing to Landy in his hotel room: "I'm a patriot. I serve my country," and calling Danny Zorn's death "collateral damage"). He knew that he would be implicated (he had sabotaged the sale of the Neski files, framed Bourne with the help of the Russian assassin Kirill, and murdered Zorn).
Jason then traveled to Moscow by train, where he was involved in an extended high-speed car chase through the crowded streets between himself (driving a hijacked yellow taxi), assassin Kirill (re-sent by Gretkov to finish Bourne off) in a stolen black Mercedes SUV, and multiple police cars ordered by the CIA, that ended in a spectacular tunnel crash. Bourne was satisfied that Kirill was about to die from his massive injuries. Gretkov was arrested by Russian police by orders from Landy, after she listened to Abbott's tape.
Bourne then sought out Irena Neski (Oksana Akinshina), living in an apartment outside of the city (in the Orannyi Projects), to apologize to her for killing her parents (and to set the record straight that her mother didn't kill her father and then commit suicide). She sobbed as he told her: "I killed them. It was my job. It was my first time. Your father was supposed to be alone. But then your mother came out of nowhere and I had to change my plan. It changes things, that knowledge, doesn't it? When what you love gets taken from you, you wanna know the truth. I'm sorry."
Then, after traveling to NYC, he telephoned Landy in her office there to hopefully end his involvement in the Treadstone debacle, and to ask her why the CIA was still looking for him ("What do you want?"). She thanked him for the tape ("We got what we needed. It's all tied off. It's over"), and then gave him an off-the-record apology. She then supplied him, from his top-secret file, with his real name (David Webb), birthdate (4-15-71) [Bourne's actual birthdate was 9-13-70] and birthplace (Nixa, Missouri). She asked: "Why don't you come in and we'll talk about it" - but he declined by peering at her in her office from afar and telling her: "Get some rest, Pam. You look tired." In the final shot, he disappeared into a crowded sidewalk in downtown Manhattan.
