At least 45 people have been killed in similar attacks in the Central American nation's capital this year.
Officials says that gang mambers are targeting bus drivers to steal takings to fund thier criminal activities.
The violence has led to the temporary closure of some bus routes.
The government said that it would provide a reward for any information which leads to the capture of Tuesday's assailants.
Gang violence in Guatemala leads to thousands of deaths every year and contributes to the nation's reputation as one of the region's most dangerous countries.