From a tactical point of view, his invasion of Russia wasn't obviously bad from the start. He couldn't launch his invasion of Britain until he had air superiority. Until he conquered the RAF, he just had ground troops with nothing to do. From his own published work Mein Kampf, you get the impression that conquering Russia primarily for resource and farming development was on his to do list eventually. From knowledge of previous wars, it would seem likely that he only signed the non-aggression pact to lull Stalin into a false sense of security and prevent fighting a war on two fronts (which up until that point he had succeeded in doing). Certainly, with the benefit of hindsight, the invasion of Russia was a mistake, but his troops did very well initially and if they had actually surrendered when he took the capital, then he would have won. Russian winter and tough citizens then decimated his army (yes, I do mean it in the reduced by 1/10 sense, but it was likely higher than that).
If one had to cite a single HUGE mistake, I would say it was signing a pact with Japan and declaring war on the US along with Japan after the Pearl Harbor bombing. Until that point, the US didn't take any official, overt action against Germany.