During a lease-signing ceremony on Thursday, Assembly Economic Development Minister Andrew Davies said the benefits the office would being to Wales would be "huge".
The high-profile business-orientated outpost is set to cost the assembly £220,000, with more taxpayers money coming from the Welsh Development Association (WDA).
"This office is about creating jobs in Wales and giving Wales a presence in North America to assist our exports, our inward investment and our tourist industry," Mr Davies said.
"To make any kind of impact, Wales needs a location at the heart of New York's business district. This new office fills that role."
But the project - the first of four across the world - has been criticised as a set of unnecessary "mini-embassies".
Assembly leaders hope the acquisition of the New York base will do much to raise Wales' profile in America.
First Minister Rhodri Morgan announced the setting up of the international centres during a St David's Day promotional trip to New York.
There had been envy over the access fellow Celtic countries Ireland and Scotland have to US business bosses and want to create a communication channel of their own, separate from that of the UK government.
It is also hoped that a presence will help foster cultural links between the US and Wales.
Other international centres are expected to open in San Francisco, Tokyo, Hong Kong and Brussels - although their cost has yet to be disclosed.
The expansion in the US and the Far East comes as the assembly is withdrawing funds from the Wales European Centre, which was set up in 1991.
The WEC is a shared facility which the assembly jointly funds with the Welsh Development Agency and the Welsh Local Government Association.
In April, the Welsh Assembly Government decided to stop its £189,000 funding of the centre and strengthen its own Brussels office instead.