Release of Cell-Free Ice Nucleators from Three Recombinant Ice+Zymomonas mobilis Strains

Abstract

<b><i>Background/Aims:</i></b> This work is a study of the ability of three recombinant <i>Zymomonas mobilis </i>strains to release ice nucleators into their growth medium. <b><i>Methods:</i></b> The recombinant ice⁺<i>Z. mobilis</i> cells were tested for their ability to produce cell-free ice nucleators, under three different growth temperatures and three different glucose concentrations. <b><i>Results:</i></b> Cell-free ice nucleators were obtained from all the recombinant ice⁺<i>Z. mobilis</i> cells tested. The cell-free ice nucleation activity was not affected by the glucose concentration in the growth medium or the growth temperature. The freezing temperature threshold was below -7.6°C, demonstrating a class C nucleating structure of the ice nucleation protein. The size of the ice nucleators was less than 0.22 μm and their density was estimated as 1.024 ± 0.004 g/ml by Percoll density centrifugation. The properties of the detected ice nucleators, in addition to the absence of pyruvate decarboxylase activity in the spent medium (a cytosolic marker), support that the cell-free ice nucleation activity was due to the extracellular release of ice nucleators. <b><i>Conclusion:</i></b> These findings indicate that the recombinant ice⁺<i>Z. mobilis</i> cells could be valuable for future use as a source of active cell-free ice nucleation protein.