All New Routemaster buses operating in the capital will be converted to make the rear and middle doors exit only.
The move follows research which showed passengers using the doors to dodge fares was costing TfL around £3.6 million a year.
In addition, a pilot on the route 8 proved using only the front doors for passenger boarding caused no delays.
Boarding using the middle and rear doors on all New Routemasters will be phased out, starting with the routes 55 and 267 on 25 January.
Customers will be advised of the changes to boarding with emails and on-board announcements. Signage will also be changed on buses. Customers with wheelchairs and pushchairs will continue to be able to board through middle doors.
TfL’s team of more than 450 revenue inspectors operate across the whole network day and night, using new technology and intelligence to identify anyone travelling without a ticket.
Fare evasion is a serious criminal offence that could lead to prosecution, a criminal record and a fine of up to £1,000. The changes being made to New Routemasters are part of a wider programme of activity designed to drive down fare evasion across the whole TfL network.
Claire Mann, TfL’s Director of Bus Operations, said: “Fare dodging is criminal and unfair, and it deprives London’s transport of the investment it needs.
“Our successful pilot on the route 8 has shown that boarding using the front door only doesn’t delay the service and reduces fare evasion, so we will change all other New Routemasters to follow suit.
“Customers with wheelchairs and pushchairs will still be able to board using the middle doors and will find it easier to access the priority space on buses as – like on other buses – they will be able to use the middle doors to board without having other customers doing so at the same time.”