Z-DNA has attracted interest due to its distinctive left-handed helical structure. This non-canonical DNA structure is able to form transiently and plays an important role in cellular processes such as transcriptional regulation and DNA recombination. Alternating purine-pyrimidine sequences are well known to form Z-DNA under high-salt conditions, but the detailed mechanism of B-to-Z transition of DNA containing BZ junctions under these conditions is not well understood. Here, using single-molecule FRET and circular dichroism experiments, we studied the effect of BZ junctions on Z-DNA formation under high-salt conditions. Further thermodynamic analysis revealed that a discrepancy of different DNA substrates in the presence and absence of BZ junctions in Z-DNA formation can be attributed mainly to the competition between enthalpy and entropy. Salt-induced B-to-Z transition is entropically favored in the presence of BZ junctions and is enthalpically favored in their absence. This thermodynamic information provides a deeper understanding of Z-DNA formation of DNA containing BZ junctions.