Bank junction will close to all vehicles apart from buses from 22 May onwards, City A.M. can exclusively reveal.
The closure of all the entrances to the junction is designed to reduce accidents at one of London's busiest crossroads. It will be enforced between 7am and 7pm, Monday to Friday, and drivers will be re-routed by advanced warning signs.
Cyclists will still be allowed through the junction, but taxis will be banned, despite repeated protests by black cab drivers.
Steve McNamara, general secretary of taxi drivers' union the Licensed Taxi Drivers' Association (LTDA), has previously described the plans as "the craziest scheme anyone has suggested for some time".
The City of London decided to exclude cabs from the junction on the basis that although they were not involved in a large number of accidents, they significantly increased congestion and therefore made the junction more dangerous.
The car ban at Bank will last for 18 months and will be kept under regular review. It is part of a wider plan to improve safety in the Square Mile, and it is not yet known how the ban will be revised after this trial period.
In the first few weeks of the new system, drivers breaking the ban will receive warning letters, but after that, automatic number-plate recognition software will be used to issue drivers with penalty notices.
The penalty for those flouting the rules will be £130, but will be reduced to £65 if paid within 14 days of the issue date.
The City of London has considered the full pedestrianisation of Bank during working hours. However, if this went ahead, it would not be implemented until 2020 at the earliest.