A suspected attacker was also killed in the ambush.
Authorities closed the road to avoid further incident in the province where the government has faced increasing secessionist protests in recent months.
The attack followed an assassination attempt on Yemens' deputy prime minister, on Thursday.
Fighters opened fire on the minister's office convoy in a different southern province, where he had attended celebrations marking the 20th anniversary of Yemen's unity.
North and south Yemen united in 1990, but many in the south - where most of the impoverished Arab country's oil facilities are - complain that northerners take their resources and deny them political rights.
Several soldiers, separatist gunmen and bystanders have been killed in the recent month's escalating tension in the south.
The government, struggling to stabilise a fractious country in which central authority is often weak, faces international pressure to quell domestic conflicts in order to focus on fighting a resurgent al-Qaeda.