The US Department of Justice is suing Barclays for alleged mortgage securities fraud.
It claims that Barclays misled investors about the quality of loans backing securities in the run-up to the financial crisis.
Barclays rejected the claims, saying they were “disconnected from the facts”.
The banking giant is among a number of European banks that have been under investigation by US authorities.
On Friday, the US said Deutsche Bank had agreed to pay $7.2bn to settle a mortgage investigation.
The US Department of Justice said: “From 2005 to 2007, Barclays personnel repeatedly misrepresented the characteristics of the loans backing securities they sold to investors throughout the world, who incurred billions of dollars in losses as a result of the fraudulent scheme.”
It said it could seek civil penalties up to the amount that Barclays gained, or that investors lost.
Federal prosecutors said that as part of the alleged scheme Barclays sold $31bn in securities.
More than half of the mortgages backing the securities defaulted, the suit alleged.
Investors included “credit unions, pension plans, charitable and religious organisations, university endowments, and financial institutions, among others” said Robert Capers, the chief prosecutor in the Eastern District of New York.
Unlike in some other cases against major institutions, the Justice Department has chosen not to seek a mutually acceptable resolution.
Barclays said: “We have an obligation to our shareholders, customers, clients, and employees to defend ourselves against unreasonable allegations and demands.
“Barclays will vigorously defend the complaint and seek its dismissal at the earliest opportunity.”
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