The Roma head coach will be a spectator for Monday’s (AEDT) game against Sassuolo and the tussle with city rival Lazio seven days later.
His ban was temporarily suspended last week, allowing him to take charge of a game against Juventus, but it now takes immediate effect.
The 60-year-old was sent off for the third time this season just two minutes into the second half of Roma’s 2-1 loss at lowly Cremonese on Wednesday 29 February (AEDT).
Mourinho clashed with fourth official Marco Serra and was sent to the stands, before continuing to argue his case when entering the officials’ dressing room after the match.
The Italian Football Association (FIGC) announced a sports court of appeal rejected Roma’s challenge to the ban, with Mourinho also fined €10,000 ($16,160) for his behaviour.
The FIGC statement said Mourinho’s punishment was imposed “for vehemently and provocatively contesting a refereeing decision, reiterating such behaviour at the time of the expulsion and also for having, at the end of the match, entered, even if authorised, the referee’s locker room and addressed to the fourth official seriously offensive expressions and inferences”.
Former Chelsea, Inter Milan and Real Madrid boss Mourinho had defended his actions after the game, saying: “I’m emotional but not crazy. For the first time in my career a referee has spoken to me in an unjustifiable way. To have the reaction I had is because something happened.”
Mourinho’s Roma side sits fourth in Serie A, ahead of last season’s champion AC Milan on goal difference.