Sequence-Based Protein Design: A Review of Using Statistical Models to Characterize Coevolutionary Traits for Developing Hybrid Proteins as Genetic Sensors

Statistical analyses of homologous protein sequences can identify amino acid residue positions that co-evolve to generate family members with different properties. Based on the hypothesis that the coevolution of residue positions is necessary for maintaining protein structure, coevolutionary traits revealed by statistical models provide insight into residue-residue interactions that are important for understanding protein mechanisms at the molecular level. With the rapid expansion of genome sequencing databases that facilitate statistical analyses, this sequence-based approach has been used to study a broad range of protein families. An emerging application of this approach is to design hybrid transcriptional regulators as modular genetic sensors for novel wiring between input signals and genetic elements to control outputs. Among many allosterically regulated regulator families, the members contain structurally conserved and functionally independent protein domains, including a DNA-binding module (DBM) for interacting with a specific genetic element and a ligand-binding module (LBM) for sensing an input signal. By hybridizing a DBM and an LBM from two different family members, a hybrid regulator can be created with a new combination of signal-detection and DNA-recognition properties not present in natural systems. In this review, we present recent advances in the development of hybrid regulators and their applications in cellular engineering, especially focusing on the use of statistical analyses for characterizing DBM-LBM interactions and hybrid regulator design. Based on these studies, we then discuss the current limitations and potential directions for enhancing the impact of this sequence-based design approach.