Nucleation of glucose isomerase protein crystals in a nonclassical disguise: The role of crystalline precursors

Protein crystallization is an astounding feat of nature. Even though proteins are large, anisotropic molecules with complex, heterogeneous surfaces, they can spontaneously group into twoand three-dimensional arrays with high precision. And yet, the biggest hurdle in this assembly process, the formation of a nucleus, is still poorly understood. In recent years, the two-step nucleation model has emerged as the consensus on the subject, but it still awaits extensive experimental verification. Here, we set out to reconstruct the nucleation pathway of the candidate protein glucose isomerase (GI), for which there have been indications that it may follow a two-step nucleation pathway under certain conditions. We find that the precursor phase present during the early stages of the reaction process is nanoscopic crystallites that have lattice symmetry equivalent to the mature crystals found at the end of a crystallization experiment. Our observations underscore the need for experimental data at a lattice-resolving resolution on other proteins so that a general picture of protein crystal nucleation can be formed.

Automated summary

Protein crystallization is a remarkable natural phenomenon, although the formation of a nucleus in this process is still poorly understood. The two-step nucleation model has emerged as a consensus, but further experimental verification is needed. By studying the nucleation pathway of glucose isomerase, it was found that nanoscopic crystallites with lattice symmetry equivalent to mature crystals are present in the early stages of the reaction process, highlighting the importance of experimental data for a general understanding of protein crystal nucleation.