CITIC Pacific and Cheung Kong (Holdings) have split up over a plan to jointly tender for the Route 3 road project.
The two companies were originally interested in joining forces to lead a consortium to compete for the franchise, estimated to be worth about $7 billion to $8 billion.
The project is a key expressway scheme involving tunnel works to serve the growing western part of Kowloon.
It is seen as a solution to the notorious traffic congestion along the Tuen Mun highway.
CITIC Pacific chairman Larry Yung Chi-kin yesterday confirmed the spilt.
He said parting with Cheung Kong was nothing more than a commercial decision but would not elaborate.
Struggle for a controlling stake in the proposed joint venture was thought to be a significant factor behind the split.