The Pueblo County Clerk and Recorder's Office stopped issuing marriage licenses to same-sex couples at 1 p.m. Monday.
The decision was made after the office reviewed a ruling that a judge handed down on Friday ordering Denver County to do the same.
Pueblo County Clerk and Recorder Gilbert "Bo" Ortiz says that a threat of legal action from the Colorado Attorney General's Office also influenced his decision.
"I stand by the initial decision I made and still believe that an individual's constitutional rights outweigh a law that discriminates against American citizens. As the Attorney General has admitted it is only a matter of time before Marriage Equality is validated by the Supreme Court of the United States," Ortiz said.
Colorado voters approved a constitutional amendment defining marriage as being between a man and a woman in 2006.
Attorney General John Suthers said last week that he has a duty to defend the law.
Pueblo County began issuing marriage licenses to same-sex couples on July 11.