While normal Spider-Man has died more times than anyone can count, the Ultimate universe ran on the rule that death was a bit more permanent, and as such not something you should just throw at a character willy nilly - and especially not for a character like Spider-Man.
After 117 issues of the web-slinger kicking ass and trading quips, the day of his death finally came at the hands of a mutated Green Goblin. After breaking out of S.H.I.E.L.D and taking the rest of Spidey's rogues gallery along with him, Osborn sets out on a murder spree to take down Peter once and for all. And with the heroes doing what they usually do in the Ultimate universe (that is to say, being dicks and fighting each other for the billionth freaking time) Spidey has to face all of his worst enemies all alone.
And as you can imagine, it does not go well for him. Though he's able to bring down Osborn once and for all, he ends up succumbing to his injuries and dying. It was a legitimately sad ending because we knew for a fact that this was the end for him, made even more clear by the introduction of Miles Morales.