Mayor Dan Jarvis and South Yorkshire council leaders have agreed to hand over a £5 million loan to the Yorkshire Wildlife Park.
The Branton-based zoo had their request granted to fund a planned expansion with the full backing of Doncaster Council at a meeting of the Sheffield City Region Combined Authority.
The park has secured additional land adjacent to the current site for a new entrance and visitor hub with themed restaurants, destination hotel and investment in signature animal reserves to expand the existing Wildlife Park by 150 acres.
SCR officers project visitor numbers will increase from 760,000 each year.
There will be also a visitor hub incorporating a 148-bedroom hotel, an events and conference venue creating more than 230 additional full time-equivalent jobs.
The money, taken from the Local Growth Fund, will be used to develop new infrastructure including access from Hurst Lane, new car parks and onsite utilities.
Wildlife park bosses may also spend the money at the request of the council to improve surrounding roads to include a roundabout at the new entrance and modifications to the Hurst Lane/Branton Road T junction.
SCR bosses predict the loan will accelerate the delivery of 315 jobs by one year and the total scheme will generate £46.2 million to the local economy each year after 10 years.
Steve Minion, chairman of Wild Life Group Ltd, said: “ The rapid growth of Yorkshire Wildlife Park and the support we have had from South Yorkshire confirmed to us that there was the potential for a game-changing project in Doncaster.
"However, turning ‘the dream’ into reality was never going to be easy and it has taken two years of intensive activity to get to this point.
"We are grateful to many partners for their help and assistance, and in particular the SCR who are providing vital support. The development will both create jobs and economic benefits to the region as well as helping add to the tourism offer in Doncaster."