Significantly different from conventional Au nanoparticles, ultrasmall Au nanoclusters (NCs) consisting of several to about a hundred Au atoms with a size below 2 nm exhibit a strong quantum confinement effect, and possess an intriguing molecular-like highest occupied molecular orbital (HOMO)-lowest unoccupied molecular orbital (LUMO) transition, quantized charging, intrinsic chirality, and special fluorescence properties, as well as high catalytic activities. In virtue of their unique molecular-like electronic structure, remarkable physicochemical properties, mild preparation conditions and good biocompatibility, Au NCs have been having a profound impact on bioanalytical and biomedical applications, such as biosensing, biological imaging, cell markers, drug delivery, photodynamic/photothermal therapy, and biomedical toxicology. As an indispensable part of Au NCs, shell ligands not only stabilize and protect the structure of Au NCs, but also have an important influence on the structure and biocatalytic activities of Au NCs. Nevertheless, the effect of shell ligands on the biocatalytic activities of Au NCs has not been paid much attention or even ignored. In this Focus article, thus, the structure and biocatalytic activities of Au NCs are discussed from the perspective of the shell ligands. Particular emphasis is directed to the discussion and exploration of the undisclosed and neglected roles of shell ligands in the biocatalytic activities of Au NCs, which are of fundamental importance to the unraveling of charge transfer behaviors and biocatalytic processes of Au NCs. In addition, the future directions to explore the mechanism of shell ligands affecting the biocatalytic activities of Au NCs, such as surface ligand engineering of Au NCs, advanced surface/interface in situ characterization techniques, theoretical analysis, and the nanobiology of Au NCs, are also put forward.