Spurs head coach Juande Ramos omitted Younes Kaboul from the squad for this game, with Didier Zokora dropping back from midfield to partner Michael Dawson in central defence.
After a slow start to the game, Aaron Lennon and Pascal Chimbonda combined well on the right and the latter's low cross forced keeper Daniel Zitka into a good claim at the near post.
Sixteen minutes in Steed Malbranque's cross from the right was just too high for Robbie Keane, before the home side started to make progress through Marcin Wasilewski on the right.
Mbo Mpenza should have done better after 23 minutes, but he miscontrolled a pass from Olivier Deschacht when he had a clear run in on goal.
On the half-hour Jermaine Jenas' free-kick was headed clear by Nicolas Frutos, who tested Paul Robinson with a low 20-yarder soon after.
Shortly before the break Robinson was down again to parry a Wasilewski drive, before getting a crucial touch to the follow-up from Frutos.
Soon after the restart Robinson had to be alert to block a Frutos effort, before Zokora made a superb recovery tackle to prevent Mpenza from opening the scoring.
Just before the hour Spurs brought on Berbatov to try and shake things up, but it was the home side who made the breakthrough midway through the second half.
Cyril Thereau's flick was allowed to run across the area by both Dawson and Jenas and Bart Goor swooped to drill a left-foot shot beyond Robinson.
Within two minutes Spurs won a penalty as Jelle Van Damme was adjudged to have brought down Jenas in the area and Berbatov coolly sent Zitka the wrong way from the spot.
Anderlecht came closest to snatching a late winner as Wasilewski's ball in from the right ran all the way across the area, with Robinson forced to scramble Thereau's poke towards goal away to safety.