Coach operator FlixBus has been served notice to stop operating from Leeds Bus Station.
The company was due to expand its services from the station next month.
FlixBus head of operations Hayley Russell said: "The decision is an insult to the people of Leeds, who are being denied access to affordable coach travel from their publicly owned bus station."
West Yorkshire Combined Authority (WYCA), which owns the station, said the move followed "a number of safety issues in recent months related to Flixbus's compliance with our conditions of use".
The coach operator has been running services in Leeds since 2021, and from the bus station since 2024.
"It's the passengers that it's going to affect more than anyone," said Russell.
She added: "We've operated there since 2024 after a few years of really pushing to be in the bus station, which is where we believe is the right place for passengers in the city."
The operator has been planning a 50% expansion in its services to Leeds due to customer demand which is set to begin on 1 April.
FlixBus maintains the plan has not changed.
Passengers hoping to travel with FlixBus will soon have to board from the roadside.
"They're being forced out of a clear place that should be designed for transport, onto a less satisfactory roadside stop," said Russell.
"We've never been banned from a station in the UK ever before."
According to FlixBus, Leeds is in its top 1% of destinations.
"Banning FlixBus from the bus station raises serious questions about why passenger choice is being restricted by WYCA, an authority led by a mayor who promised integrated transport and better buses to the region," Russell said.
FlixBus estimates that annually in West Yorkshire coaches alone contribute more than £425m to the local economy.
A WYCA spokesperson said:
"We treat all bus and coach companies equally, with regular performance and compliance monitoring of each operator using our facilities to ensure they operate with safety as their primary focus.
"Unfortunately, a number of safety issues have arisen in recent months at Leeds Bus Station related to Flixbus's compliance with our conditions of use, which has led us to serve notice on their usage of the bus station.
"We will now discuss with them how and when we best transfer their existing services to a suitable alternative location.
"Our priority as always is the safety of our staff and passengers using our bus stations."