The fixation of the catalyst interface is an important consideration for the design of practical applications. However, the electronic structure of MoS2 is sensitive to its embedding environment, and the catalytic performance of MoS2 catalysts may be altered significantly by the type of binding agents and interfacial structure. Interfacial engineering is an effective method for designing efficient catalysts, arising from the close contact between different components, which facilitates charge transfer and strong electronic interactions. Here, we have developed a layer-by-layer (LbL) strategy for the preparation of interfacial MoS2-based catalyst structures with two types of conducting polymers on various substrates. We demonstrate how the assembled partners in the LbL structure can significantly impact the electronic structures in MoS2. As the number of bilayers grows, using polypyrrole as a binder remarkably increases the catalytic efficacy as compared to using polyaniline. On the one hand, the ratio of S22- (or S2-), which is related to the remaining active hydrogen evolution reaction (HER) species, is further increased. On the other hand, density functional theory calculations indicate that the interfacial charge transport from the conducting polymers to MoS2 may boost the HER activity of the interfacial structure of the conducting polymer/MoS2 by decreasing the adsorption free energy of the intermediate H* at the S sites in the basal plane of MoS2. The optimized catalytic efficacy of the (conducting polymer/MoS2)n assembly peaks is obtained with 16 assembly cycles. In preparing interfacial catalytic structures, the LbL-based strategy exhibits several key advantages, including the flexibility of choosing assembly partners, the ability to fine-tune the structures with precision at the nanometer scale, and planar homogeneity at the centimeter scale. We expect that this LbL-based catalyst immobilization strategy will contribute to the fundamental understanding of the scalability and control of highly efficient electrocatalysts at the interface for practical applications.