I don't normally use ISO files, so I'm just speculating. However, as I understand it, an ISO file is a disk image that includes everything that would go on an optical disc, including the file system overhead. Blu-rays use the
UDF file system, usually version 2.5. I don't know what the overhead is for UDF, but it is not zero.
When you use MakeMKV to create a .mkv file or a backup of the whole disc, it does not contain any UDF overhead.
I would guess this filesystem overhead accounts for the different in size between an ISO of a disc and a MakeMKV backup.