3D Structure Prediction and Structural Stability Profiling of Cub-Mutated iRhom2

Abstract

The iRhoms are part of the rhomboid family, highly conserved among all sequenced metazoans. However, they are known as pseudoproteases as they lack vital amino acid residues required for the catalysis of serine proteases. The iRhom2 consists of a large cytoplasmic N-terminus and an inactive catalytic domain of an unknown function. iRhom2 is an unstable protein, which performs a crucial part in regulating the EGFR pathway by the maturation of TNF-α converting enzyme, TACE or ADAM-17. The up and downregulation of the EGFR pathway by changing iRhom2 can lead to several human diseases, including breast cancer, Alzheimer's disease, TOC and many others. The stability of iRhom2 can be increased via some specific mutations induced at the N-terminus. The deletion of 268 amino acids from the N-terminus results in a gain of function mutation in iRhom2; this mutation is known as cub-mutation, observed in mice. The cub-mutated iRhom2 can hyperactivate or downregulate the EGFR pathway. The hyperactivation leads to increased wound healing, inflammation and enhanced tumorigenesis, thus making this pseudoprotease more stable than wild-type iRhom2. However, the downregulation of the EGFR pathway cannot make cub-mutated iRhom2 stable than wild-type iRhom2. The contradiction about the stability of cub-mutated iRhom2 is investigated in this study. The 3D protein structure prediction, protein structure evaluation and MD simulation techniques were used to solve this contradiction. Ab-initio protein modelling technique was used to predict cub-domain (1-268) and complete cytoplasmic N-terminus domain (1-351) of iRhom2 for comparative analysis of both domains. The evaluations of these predicted domains' structures by various evaluation methods result in possible accurate models. In addition, the MD simulation of both iRhom2 domains was completed to check the structural accuracy and stability of these domains. The MD simulation results showed that the complete cytoplasmic N-terminus domain is modelled as a less compact and unstable structure. However, the cub-domain of iRhom2 resulted in a more stable and compact structure than the complete cytoplasmic N-terminus domain. The contradiction about the stability of cub-mutated iRhom2 is resolved; however, extensive insilico and in-vivo research are required to explore the functional stability of iRhom2 further.