Israel international Kayal was sidelined for four months with ankle ligament damage after a tackle by McCulloch during Celtic's 1-0 win against Rangers in December 2011.
In an interview with Israeli newspaper Haaretz, Kayal spoke about the challenge and said: "The injury didn't look that bad but I believe he did it on purpose. That player is not the cleanest in the world."
But McCoist, speaking ahead of the club's Ramsdens Cup Final against Raith Rovers on Sunday, hit back at the accusations about his skipper.
"Anybody who knows Lee, knows he is not a dirty player," the Rangers manager said.
"I don't think he is a dirty player at all. I can't even remember the tackle or the incident."
McCoist, who played for Rangers for 15 years between 1983 and 1998, also added that the Old Firm games of recent years were tame in comparison to previous encounters.
"With the greatest respect to Kayal, I would have the Old Firm games down as quite tame in comparison to some of the previous Old Firm games," he said.
"I am not sure he would have enjoyed playing with Bertie Auld (former Celtic midfielder) and John Greig (former Rangers captain) in some of those Old Firm games, that would maybe have been a wee eye-opener for him.
"But as I say, I would need to defend my player. He is not a dirty player at all and we are delighted with the skipper, the way he handles himself on and off the park."
A win against Raith Rovers at Easter Road on Sunday (April 6) would give Rangers their second piece of silverware this season.
They secured the Scottish League One title and promotion to the Scottish Championship with eight games of the season still remaining when they beat Airdrieonians 3-0. McCulloch scored all three goals on that occasion.
The Glasgow club are also in the last four of the Scottish Cup and will face Dundee United at Ibrox for a place in the final.