Unfolding mechanism of proteins is an old and new problem. In this article, basic mechanism of protein unfolding are explained using the examples of pressure-induced unfolding, denaturant-induced unfolding, and thermal and cold unfolding. Although unfolding of small globular proteins follow two-states, native and unfolded states, it does not mean the existence of only two structures, since the states are determined by intermolecular interactions but the structures are determined by intramolecular ones. Pressure-induced unfolding clarifies important contribution of hydration to the unfolding mechanism of proteins, because pressure affects to total volume of the system. Denaturant-induced unfolding are explained by preferential interactions between protein and denaturant molecules, which mechanism can also explain effects of stabilizers such as salts or sugars although sign of the interactions are inverted. Negative entropy change due to cold unfolding indicates extensive restriction of hydrated water molecules exceed to the increase of protein’s conformational chain entropy due to unfolding. Contrary to instinctive images, thermodynamic parameters showed that the folding of several proteins at room temperature raise the randomness of protein solution in the case of cytochrome c, myoglobin, tryptophan synthase α subunit, and human serum albumin.