I know Reg E does not apply to business accounts, so what is the general practice on paying and charging for ATM and one time debit card transactions on those accounts?
Since in our system the Reg E Opt-In status must be marked with something, even on business accounts, we just have it marked as "decline", and do not generally allow those charges to come through at this point. Unless, of course, it happens on accident, and then we don't charge a fee.
How's everyone else handling these transactions?

I don't recommend allowing a business to "opt-in" to an overdraft protection "privilege" program, whether it affects card transactions or not. Although I understand there is no commitment to extend credit under a Privilege program, approval of an overdraft is an extension of credit, and many of the individuals with business account-connected debit cards do not have authority from the business to borrow.