The independent commission, comprised of former prosecutors from outside Guatemala, will have an initial two-year mandate to gather evidence and help build cases against illicit criminal groups. Suspects would be tried in local courts.
Some organized crime groups are believed to be made up of former military members who fought in Guatemala's 36-year civil war that left about 200,000 dead.
Peace accords ended the fighting nearly 10 years ago, but violence and corruption continue. The U.N. has said about 5,000 murders are committed annually in Guatemala, and U.S. officials estimate that three-quarters of all cocaine consumed in the U.S. passes through the country, a key smuggling point between Colombia and Mexico.
''With this agreement, the United Nations is standing by Guatemala as it tries to solidify democracy and the rule of law by exposing and dismantling criminal groups that grew out of the armed conflict,'' Ibrahim Gambari, the U.N. undersecretary-general for political affairs, said in a statement.
Guatemalan Vice President Eduardo Stein, who signed the agreement with Gambari at U.N. headquarters on Tuesday, said earlier that the integrity of the commission would be guaranteed by prosecutors with ''vast experience investigating criminal networks.''
The commission still must be approved by Guatemala's Congress.
Authorities hope the commission will help beef up the country's criminal justice system, which has been targeted recently by organized crime groups.
The Washington Office on Latin America, a human rights advocacy group, said in a statement Tuesday that criminal elements have terrorized judges, witnesses, prosecutors and human rights defenders in Guatemala over the last several years. The group said six justice officials were assassinated last year and five politicians were killed this year.
A U.N. human rights observer said in August that most violent crime in the country goes unpunished because of lack of investigation.
''It's sad to say, Guatemala is a good place in which to commit a murder. Your chances of being committed and punished are staggeringly low,'' said Philip Alston, the U.N.'s special investigator on extrajudicial, summary or arbitrary executions.
Associated Press Writer Juan Carlos Llorca contributed to this report from Guatemala City.