- In 16th-century Prague, a rabbi creates the Golem - a giant creature made of clay. Using sorcery, he brings the creature to life in order to protect the Jews of Prague from persecution.
- Rabbi Loew, the leader of medieval Prague's Jewish population, studies the stars and warns the local elders of impending doom for his people. Squire Florian delivers the seeming fulfillment of that prediction by delivering him a proclamation from the Holy Roman Emperor that orders the Jews to evacuate the city before the new moon. Meanwhile, Loew starts to come up with a strategy for defending the Jews. In his visit, the conceited Florian is drawn to Miriam, Loew's daughter. His rival for her affections is Loew's assistant. Loew persuades Florian to remind the Emperor that he has told the Emperor's horoscopes and prophesied tragedies, and he asks for an audience. Loew starts to create the Golem, a gigantic clay creature that he would give life to protect his people. Later, Florian arrives bearing a letter from the Emperor inviting Loew to the palace's Rose Festival.
Loew tells his assistant that he has secretly constructed the Golem and gets his help animating it. Loew and the assistant use a complex magical ritual to call forth Astaroth the spirit and force him to speak the word of life, as prescribed in the old books. Loew inscribes the word on paper and secures it inside an amulet fastened to the Golem's chest. Golem awakens and the Rabbi initially employs it as a domestic servant.
Loew brings the Golem with him to the palace's Rose Festival to impress the audience. Florian slips away from the court to meet Miriam, whose house is being guarded by Loew's assistant. Back at the palace, the court is both terrified and intrigued by the arrival of the Golem. Impressed, the Emperor asks to see more supernatural feats. Loew projects a magical screen showing the history of the Jews, instructing his audience not to laugh or even speak. The court starts to chuckle when Ahasuerus, the Wandering Jew, arrives, and the palace suddenly starts to collapse. Loew commands the Golem to respond and he supports the crumbling ceiling, saving the court. The Jews are forgiven and let to stay by the Emperor as a token of gratitude.
Loew and the Golem return to the ghetto and Loew shares the news that the Jews are saved. Loew returns to his house and begins to notice erratic behavior in the Golem. After managing to remove the amulet, he reads that upcoming astrological movements will cause Astaroth to possess the Golem and attack its creators.
While the neighborhood is jubilant, the assistant rushes to tell Miriam but discovers her in bed with Florian. Devastated, he puts the amulet back in the Golem again and commands it to remove Florian from the home. However, the Golem, now under Astaroth's control, violently tosses Florian off the roof of the home, killing him. Miriam faints when she sees what the Golem has done. The Golem carries her back down into the house. When the assistant sees the Golem's affection for her, he tries to take the amulet out of the Golem's chest. Enraged, the Golem goes after the assistant by swinging a lit log from the fireplace at him and inadvertently setting the house afire. The Golem drags the unconscious Miriran by the hair down to the street.
Loew's assistant rushes to the synagogue to alert the praying Jews of the disaster, but upon their arrival at Loew's house, they find that it is burning, and both the Golem and Miriam are missing. Despaired, the community begs Loew to stop the fire. Loew performs a incantation that stops the fire but also removes Astaroth from the Golem. Promptly, the Golem, who is wandering the ghetto causing destruction, leaves Miriam. Loew finds Miriam and she awakens. Loew's assistant joins them and tells Loew that the Jews are waiting for him by the gate. After Loew has departed, the assistant begs for forgiveness from Miriam and swears to never tell anybody about her illicit relationship with Florian.
Golem moves in the direction of the city gate, opens it and spots some young girls playing. He scoops up one unafraid girl while all the others run away. Curious, she takes the amulet out of the Golem. He drops her and falls backward, lifeless. The little girl then runs to find her friends again.
Loew makes for the gate but encounters the Jews on the way who are looking for the Golem.
The little girl returns with all the other girls to where the fallen Golem lays. Now unafraid, they sit on his still body. The little girl tosses the amulet away.
Loew and the other Jews are told the Golem has opened the gate. The crowd rushes to the gate and find the girls sitting on the Golem's body. The girls run away in fear. Loew gives praise to Jehovah for saving them a third time that day and all the Jews briefly bow down in worship of Jehovah. They then get back up and lift the Golem up and take him back into the ghetto, rejoicing and praying.
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