Understanding the nature of the interaction between small molecules and G-quadruplex DNA is crucial for the development of novel anticancer drugs. In this paper, we present the first data on time-resolved fluorescence anisotropy study on the interaction between a water-soluble cationic porphyrin H2TMPyP4 and four distinct G-quadruplex DNAs, that is, AG3(T2AG3)3, thrombin-binding aptamer (TBA), (G4T4G4)2, and (TG4T)4. The anisotropy decay curves show the monoexponential for free H2TMPyP4 and the biexponential upon binding to the excess amount of G-quadruplex DNAs. The biexponential anisotropy decay can be well interpreted using a wobbling-in-the-cone model. The orientational diffusion of the bound H2TMPyP4 is initially restricted to a limited cone angle within the G-quadruplex DNAs, and then an overall orientational relaxation of the G-quadruplex DNA-H2TMPyP4 complexes occurs in a longer time scale. It was found that the dynamics of the restricted internal rotation of bound H2TMPyP4 strongly depends on the ending structures of the G-quadruplex DNAs. According to the order parameter (Q) calculated from the wobbling-in-the-cone model, we deduce that the degree of restriction around the bound H2TMPyP4 follows the order of TBA>(TG4T)4>AG3(T2AG3)3>(G4T4G4)2. Especially, based on the maximum order parameter (Q) of bound H2TMPyP4 within TBA, a new sandwich-type binding mode for TBA-H2TMPyP4 complex was proposed in which both terminal G-quartet and T•T base pair stack on the porphyrin ring through π-π interaction. This study thus provides a new insight into the interaction between G-quadruplex DNAs and H2TMPyP4.