The statement from the presidency said Paillat's resignation had been accepted by President Ali Bongo Ondimba, without giving further details.
The deputy transport minister will take over his responsibilities until a replacement is appointed, the statement added.
There had been growing calls for the departure of Brice Paillat from relatives of the victims, opposition politicians and campaigning groups since the disaster off the coast of Gabon.
The government had already suspended four senior figures inside the merchant marine and maritime affairs administration.
The Esther Miracle ferry, carrying 161 passengers and crew, went down early on Thursday, March 9, during an overnight voyage from the capital Libreville to the oil town of Port-Gentil.
The toll, while still not definitive, has remained the same for the last two days.
The ferry had been bought and launched on that route last November and belonged to a private company, Royal Cost Marine (RCM). It is not clear when they vessel was built.
Witness accounts reported by various media said the crew did not help passengers as they tried to evacuate the sinking boat.
Many were left clinging to inflatable rafts as they took on water for hours, before the first help arrived. The ferry sank 10 kilometers off the coast.
Campaigners and relatives of the dead have questioned why the vessel, which was in a poor condition, was being used to carry both freight and passengers.
The authorities have opened two investigations into the disaster, but so far, prosecutors have said only that some people have been detained.
