Accompanied by television cameras, Putin set out on a 2-thousand kilometre (1,242 mile) drive from Khabarovsk towards Chita in Siberia, previously only reachable by rail or air.
Talking to reporters before the start of his journey, the premier said the route has historic significance - providing a vital link between western and eastern Russia.
"Our country has the biggest territory in the world and despite that it never had a full highway connection," he said before getting into his yellow Russian-produced Lada Kalina Sport car.
He said he'll be observing the results of the long-running project from the driver's seat, and inspecting key construction sites on the way.
Putin was filmed stopping at a petrol station and having a snack with locals, discussing the rise in bread prices with them.
Among the key sites Putin was to visit during his journey was a hydroelectric plant, one of the biggest in the country, and a soon-to-be constructed space launch pad, Russia's second one.