As someone who grew up watching many Disney classics, I couldn't not pick Ursula, Maleficent and Cruella. I'd also add the Queen and Lady Tremaine to the list.
And personally I would put O-Ren Ishii over Elle Driver any day of the week, if only because of that awesome anime sequence about her past.
However, here's a few other favorites of mine:
Baby Jane Hudson (
What Ever Happened to Baby Jane?)
She beats up her disabled sister and murders innocent parrots, but is she really the villain?
Eve Harrington (
All About Eve)
The archetypal snake in the grass. So devious, she's even bisexual.
Kathie Moffat (
Out of the Past)
One of the best femme fatales ever. Poor Robert Mitchum [blackout]had to get himself killed in order to finally stop her[/blackout].
Vermithrax Pejorative
Hey, she's a lady too, and the best movie dragon to date. Also, how many monsters can claim to have actually managed to eat the princess? Well, she can.