We have firstly demonstrated the renaturation process of dissociated single chains of lentinan (s-LNT) and the variable conformations of the renatured LNT (r-LNT). The results from ultrasensitive differential scanning calorimetry and circular dichroism revealed that the variable structures including perfect triple helix, defective triple helix containing duplex segment, and single chains occurred in the renaturation of s-LNT, depending on the renaturation time, solvent composition, molecular weight, and the mode of renaturation. When water was added into s-LNT/dimethylsulfoxide (DMSO) to reach 95% (v/v), the classic low-temperature intra-triple-helical conformational transition at ∼10(°) C (T(1)) appeared within 4 h, indicative of a rapid reconstruction of triple helical structure. Besides, one newly endothermic peak at ∼43(°) C (T(2)) simultaneously occurred, which was first ascribed to the melting of duplex segment in the imperfect triplex. The duplex stretches disappeared when DMSO reached 50%, in which single chains coexisted with triplex. Moreover, the duplex segment disappeared by slowly dropping water into s-LNT/DMSO. This work suggested that the structure of r-LNT could be controllable, and provided important information for their successful development and application in polymer and life science.