The yeast surface display system is a valuable platform for constructing cells with novel functions for various applications and high-throughput screening of protein or peptide libraries containing random mutations. Among the host microorganisms used for surface display, yeast is the most suitable microorganism for surface engineering owing to its eukaryotic features. In yeast, proper folding and glycosylation of expressed eukaryotic proteins can be performed. Furthermore, in this system, multiple kinds of proteins can be simultaneously displayed on the cell surface. This allows for a synergistic effect between the displayed enzymes, leading to an efficient multistep reaction. Alternatively, the ratio of the enzymes to be displayed can be controlled by the co-culture of surface-engineered yeasts displaying a single kind of enzyme. Therefore, yeast surface display systems have been applied to the construction of various whole-cell biocatalysts. Here, we describe methods for the simultaneous display of multiple kinds of proteins on the yeast cell surface.