The Turkish raids of 1471 and especially of 1498 were devastating for the eastern coast of Crete, after the settlements and monasteries of the area were looted and of course the Monastery of Akrotiriani which held the prominent position of the monastic community. The rebuilding of the new Monastery in place of the old one was related to its precarious position, which is why it took the necessary defensive-fortress form to deal with the Turkish danger. The rebuilding of the Monastery in a fortress form was associated with the Venetian Cretan families of the Kornaris (e ioia) and the Metzos (Moooa) of Sitia, which is why even today the south wing bears the name of the Kornaris and the north of the Metzos, names that witness the sponsors of the Monastery.

The Monastery flourished in the 14th and 15th centuries, judging by the large number of important Byzantine icons from that period. The images faithfully echo the evolution of Constantinople painting that gradually entered Crete from the fall of the city onwards. The high artistic value of the images is also indicative of the high level of education of the monastic community of the Monastery, which played an important role in the rise of the cultural level of Renaissance Crete. This period of prosperity of the Monastery and of the Renaissance Cretan culture in general continued undisturbed throughout the 16th century, until 1612, when its smooth course was interrupted by the devastating earthquake that hit Eastern Crete.

The Venetian Senate, considering the Monastery a stronghold for the defense of Eastern Crete, significantly strengthened the restoration of the earthquake damage. The work was undertaken by the learned and active Abbot Gabriel Pantogalos, who was the scion of an urban family of Sitia. Gabriel chose the architectural type of the Monastery and implemented it, giving the current impressive form of the fortress complex, supported by the Venetian nobleman Andreas Cornaros. The restoration of the damage to the building complex was done with the financial resources of the Monastery, subscriptions of the faithful and with the help of the 200 ducats of the Venetian Senate. This renovation marked the beginning of a new period in the history of the Monastery. Historical evidence confirms the great prosperity of the Monastery with an increase in monks and service staff, charged with the cultivation of the rich lands with which the Monastery was endowed for the fulfillment of its military-defense mission in the region. This wealth was preserved and increased with his skilful management by Gabriel Pantogalos, who remained Abbot until 1641, when he was transferred as Abbot to Agarathos Monastery in Heraklion. His name is carved on an inscription that is built into the outer west side of the temple.