I do this trip regularly, via Highway 401 (about 330 km / 4 hours). For my little boat, it's cheaper and safer (and faster) this way.
From Trenton onward, the
route is sufficiently sheltered that anything over 20 feet or so should have no trouble in good weather. West of Trenton is pretty exposed. A boat capable of open-water passages could go right down the middle of the lake, but I wouldn't do that in a
small boat or if there's a risk of weather. There are places to put in for fuel and shelter every 50-75 km or so along the shore. Expect to be ripped off at the marina pumps (just like the Kingston gas stop with the sign saying "Last fuel on 401 in Ontario", the prices are often inflated by 10 to 15 cents a litre.)
If you decide to do it by truck and trailer, there is a free (but poorly maintained) municipal
ramp in downtown Kingston and a pricier ($10-$12) but very well kept
ramp at Portsmouth Harbour that can easily handle
boats of 30+ feet. The inner harbour marina has facilities to launch larger
boats and, from what I've seen, isn't unreasonably pricey.
Stopping near Brighton for a night lets you break the trip up into two legs done at slower speeds, and could save a fair bit of fuel
money.