ATP-binding cassette (ABC) transporters are members of a protein superfamily that is one of the largest and most ancient families with great variety of functions. They are involved in transporting different substrates across the plasma membrane. As more ABC transporters were identified from the gene clusters for antibiotic biosynthesis, recent studies have begun to focus on their biological functions. The polyene antibiotics represent a class of biologically active fungal metabolites synthetized by the genus Streptomyces. As drug resistance to polyene remains rare, polyene antibiotics have been the cornerstone of therapy for critically ill patients with invasive fungal infections. We reviewed the latest research progresses on ABC transporters of polyene antibiotic biosynthesis gene clusters, including their structure characteristics and biological functions. The potential relationship between the secondary structure and their function is also discussed.