Reverse micellar solutions aided permeabilization of baker's yeast

The yeast cell wall is highly resistant towards chemical permeabilization. In the present work, reverse micellar solutions (RMSs) of sodium di (2-ethylhexyl) sulfosuccinate (AOT) in aliphatic hydrocarbons along with aqueous solutions of various surfactants, hydrotropes and amino acids have been examined for their ability to permeabilize baker's yeast cells. Although a substantial amount of cellular proteins was released on treatment with RMSs, higher molecular weight proteins such as invertase (INV), alcohol dehydrogenase (ADH) and alkaline phosphatase (AP) were retained within the cell. Treatment of intact yeast cells with aqueous cysteine solutions not only resulted in specific release of invertase but it also made the cells more fragile, enhancing the release of proteins/enzymes in the subsequent cell permeabilization steps. A multi-step procedure comprising of cysteine treatment as the first step (a purification factor of 40 for invertase), followed by reverse micellar solid–liquid extraction (RME) removed most of the proteins from the cells. Finally, sonication released the intracellular enzymes ADH and AP in a comparatively purer form with ∼90–95% recovery.