Despite the unsuccessful attempts to obtain a crystal structure of MicA and the absence of structural data of homologous proteins, we managed to generate de novo predicted models of MicA using the tFold , RoseTTAFold , and AlphaFold 55 computational methods (Extended Data Fig. ). Among these models, the one derived from tFold ranked first according to the "Protein Model Quality Assessment" system embedded in the tFold platform (Supplementary Figure ). Consequently, we used this tFold-derived model for further analysis. The overall helical structure bore a resemblance to a classical type I terpene synthase fold, and the structural alignments with Bnd4 (predicted model) , AlbS (predicted model) , and -humulene synthase (AsR6, PDB ID: 7OC5) resulted in a root-mean-square deviation (RMSD) of 2.20 Å, the pyrophosphate moiety, also shows a significant effect (Fig. ). Furthermore, residues forming the active site cavity and interacting with carbons of GGPP, such as S187, Y152, W313, W306, M74, V220, L221, G182, G183, M184, A78, W77, F51, E53, and L54, are mainly shown in Fig. . Notably, among these residues, the indole nitrogen of W77 displays a 3.82 Å distance to C17 of GGPP (Fig. ), suggesting a probable deprotonation effect. Subsequent alanine scanning-based mutations on the above residues demonstrated their importance in eunicellane scaffold formation.