The crash between a Ford Ka and a Ford Focus happened on the A1 near Torness power station in Dunbar, East Lothian, at about 7.40pm on Thursday.
A 21-year-old woman who was a passenger in the Ka died at the scene.
Three people were airlifted to Edinburgh Royal Infirmary by a search and rescue helicopter from RAF Boulmer, while another person was taken to hospital by ambulance.
The 21-year-old woman who was driving the Ka has now died, police said.
Another female passenger, aged 20, in the Ka is being treated for injuries, along with the 23-year-old man who was driving the Focus and his 20-year-old female passenger.
The road was closed for about six hours while police carried out investigations.
Police Scotland have reissued an appeal for anyone with information to contact them on the non emergency 101 number, or anonymously via Crimestoppers on 0800 555 111.