"It's a decision that has been made ... it is not going to be reversed," Sandra Jovel told journalists during an event to commemorate the end of the Guatemalan civil war in 1996.
"The Guatemalan government is very respectful of the positions that other countries have taken, and as we are respectful of those decisions, we believe others should respect decisions made by Guatemala," she added in response to critics including the Palestinians.
The announcement made Guatemala the first country to follow the United States's controversial lead on the holy city.
Israel claims all of Jerusalem as its capital, while Palestinians, consider east Jerusalem as the occupied capital of their future state.
Morales, defending his decision, said Israel is an "ally" and that Guatemala has "historically been pro-Israel.