I'd have to make mine a top 4...
1) Doc Hopper, obviously. He was a dangerous villain that wanted Kermit so bad, he wouldn't mind having him dead. Pretty dark stuff in an otherwise wacky film. Even darker than Mel Brooks as a Nazi scientist.
2) Long John Silver. Mainly for the fact that Tim Curry hammed it the heck up and had a lot of fun in the film. Shame he hasn't worked with the Muppets since other than voice over for that Sesame Street Christmas Carol and the Milk ads.
3) Constantine. He became my favorite new Muppet character in short time. I really hope he doesn't disappear with MMW
4) Who doesn't love Nicky Holiday? May not have been their most dangerous villain, but a slimeball none the less.
As for the other villains...
I'd call Dr. Singer somewhat of a Knights Templar antagonist over a villain. he wasn't so much evil, but obsessed. Still, if there was a fifth entry from me, he'd be on it. His interactions with Gonzo and Bobo are some of the best things that film had to offer.
Tex Richman's very good if you count the full version of his story. That level of vindictiveness makes the villain more rounded out than the petty oilman that just wanted land.
And as much as Rachel Bitterman was ably performed by Joan Cussack, I just don't think she was that much of an actual threat. Her motives were no more than wanting a deed to expire to get land. Sure, she was nasty and cheated a bit, but not so much evil as a greedy jerkwad.
Scrooge does not count. He wasn't a villain, he was his own worst enemy.
And, for the sake of the subject...
The Sleaze Brothers were pretty bad, though more like greedy jerks than actual threatening evil. While Mrs. Finch is an antagonist. Her motives weren't actually bad, much less evil. She thought she was acting towards the greater good for Big Bird, making her more of a Knights Templar figure.
And of course, Huxley. A funny, affable type that doesn't really manage to make anything more than a standard preschool show villain. His "evil" is based on bad behavior in younger kids, which is used to make a bad example and to not scare the crap out of the little ones watching.