The site on Mount Zion, known as the Cenacle or Upper Room, is located in a two-storey building also considered holy to Jews and Muslims, who regard it as the site where the biblical figure David was buried.
Under Israeli law, Christians are only allowed to pray there twice a year, prompting efforts by the Vatican to negotiate greater access rights to what is one of the most sacred sites in Christendom.
Those attempts have sparked a major backlash by nationalist Jewish groups, many of whom wrongly believe Israel is poised to sign over sovereign rights to the site.
In recent years, the compound has become a rallying point for Israeli hardliners. Last year, suspected Jewish extremists smashed hundreds of 17th century Ottoman tiles off the walls at the entrance to and inside the tomb.
Tensions remain high ahead of the visit, after police arrested 26 Jewish hardliners protesting near the site on Sunday, just hours before Francis touched down in Jerusalem.
For Jews, it is the ground floor which is sacred, revered since the 12th century as the burial place of David, who ruled from Jerusalem, although the site has never been excavated and the contents of its sarcophagus are unknown.
The pope will celebrate mass at the Upper Room hours before the end of his three-day visit to the Holy Land, in a move which has sparked protests and several incidences of anti-Christian vandalism.
"All the rumours about transferring sovereignty or ownership rights of any kind are false rumours," said Yigal Palmor, spokesman for Israel's foreign ministry, dismissing them as "conspiracy theories spread by parties with vested interests."
- Freedom of access, not worship -
Up a flight of stairs is a long room, with stone pillars supporting a beautiful vaulted ceiling.
It is here that Christians believe Jesus ate the passover meal with his disciples before he was crucified, a meeting known today as the Last Supper. It is also the place where his followers were baptised by the Holy Spirit during the Jewish Feast of Weeks, known to Christians as Pentecost.
"For Christians, Mount Zion is the second place in importance after the Holy Sepulchre," says Father Pierbattista Pizzaballa, Custodian of the Holy Land, referring to the Jerusalem church built over the site where tradition says Jesus was crucified, buried and rose again.
Under an Ottoman law still enforced by Israel, the Catholic Church can only celebrate mass there twice a year - on Holy Thursday just before Easter, and on Pentecost.
"Free access is one thing, but freedom of worship is something else. If I want to go there I can, but if I want to pray there officially, as a Christian, I cannot do this," Pizzaballa said.
The Vatican has no interest in taking over the site, but only gain greater access to use the room for worship, he said.
"We don't want to transform the Upper Room into a church, we don't want property, we don't want sovereignty. We want the principle that, since it is the second most important holy place for Christians, we have the right to pray there."
- Centuries of dispute -
Mount Zion, which lies just outside the walls of the Old City, is unique in that it is holy to Jews, Christians and Muslims, all of whom have ruled over the site there at different points in history.
The compound itself, which is located in Israel territory, has two separate entrances, both leading from an outside courtyard presided over by a giant gold statue of King David.
In the first century, a small Christian community of Jewish origin lived there, and later a Byzantine church was built on the site. After that was destroyed, the Crusaders began building a new church in the 12th century, which included the tomb and the Upper Room.
Following disputes over ownership between Christians and Jews, the site was taken over in the mid 16th century by the Ottoman Sultan who declared it to be the Muslim holy site of Nabi Daud.
It was then turned into a mosque and handed over to the custody of the Dajani family who restricted Jewish and Christian prayer at the site, and held it until the newly-established state of Israel took over in 1948.
The site is under Israeli sovereignty, and all three faiths are still represented there, although it is an uneasy coexistence.
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