Human Hippo signaling pathway plays an important role in the tumorigenesis of diverse cancers and has been recognized as an attractive therapeutic target of gastric cancer. The transcriptional coactivators Yes-associated protein (YAP)/transcriptional co-activator with PDZ-binding motif (TAZ) are the major downstream effectors of Hippo, which interact primarily with transcriptional enhanced associate domain (TEAD) protein through their TEAD-binding domain (TBD). Competitive disruption of the TEAD-YAP/TAZ interaction using peptide inhibitors has been exploited as a potential strategy to treat gastric cancer by regulating Hippo signaling. Here, the crystal structures of TEAD complex with YAP/TAZ TBD domain are investigated systematically at structural, energetic and dynamic levels, from which two binding hotspots are identified; they separately correspond to an alpha-helix and a omega-loop of TBD domain, and contribute essentially to the complex interaction. Several linear peptide segments derived from the hotspot regions are highly flexibility and exhibit moderate or modest affinity for TEAD. The omega-loop-derived peptides are found to have a higher affinity, which are cyclized by introducing a disulfide bridge across their two termini. Affinity assay confirms that the cyclization can considerably improve peptide affinity by 3.7-6.6-fold. Computational analysis reveals that the designed cyclic peptides exhibit a decreased flexibility and intrinsic disorder; they can roughly maintain in native active conformation out of TBD protein context, with a reduced entropy cost upon binding to TEAD.