The role of alpha-synuclein (αS) amyloid fibrillation has been recognized in various neurological diseases including Parkinson's Disease (PD). In early stages, fibrillation occurs by the structural transition from helix to extended states in monomeric αS followed by the formation of beta-sheets. This alpha-helix to beta-sheet transition (αβT) speeds up the formation of amyloid fibrils through the formation of unstable and temporary configurations of the αS. In this study, the most important regions that act as initiating nuclei and make unstable the initial configuration were identified based on sequence and structural information. In this regard, a Targeted Molecular Dynamics (TMD) simulation was employed using explicit solvent models under physiological conditions. Identified regions are those that are in the early steps of structural opening. The trajectory was clustered the structures characterized the intermediate states. The findings of this study would help us to better understanding of the mechanism of amyloid fibril formation.