The horses - both weighing just under a tonne - have a long week ahead of them as they work their way along the river-facing meadow, which is towered over by King's College chapel.
College bosses ended a 300-year tradition of neatly mowing the grass when they transformed part of the slick lawn into a wildflower meadow last year.
This marks the first time ever that Shires have been used to trim the meadow as it reaches the end of its second year of flowering.
Last year, gardeners cut the meadow - which is home to 130 different types of insects and five bat species - by hand.
The Shires started their work just after 9am yesterday (July 2) and are expected to finish the work on Friday.
King's College's head gardener Steven Coghill said: "We’re absolutely thrilled to be bringing in these magnificent heavy horses to harvest the wildflower meadow.
"Not only do they have a far lower carbon footprint than using a rotary mower, the sight of these wonderful creatures at work in the College should make for a remarkable, bucolic scene and bring a bit of Constable to Cambridge. ”
Prof Michael Proctor, provost of King’s College said: “The wildflower meadow has inspired us to think more about how we look after and enhance our biodiversity, and has brought to attention just how much has been lost.
"We hope its presence in this iconic setting will motivate others to think about sustainability and to consider planting their own meadows elsewhere.”