Release of cell-free ice nuclei from Halomonas elongata expressing the icenucleation gene inaZ of Pseudomonas syringae

Release of ice nuclei in the growth medium of recombinant Halomonas elongat a cells expressing the inaZ gene of Pseudomonas syringae was studied in an attempt to produce cell-free active ice nuclei for biotechnological applica tions. Cell-free ice nuclei were not retained by cellulose acetate filters of 0.2 mum pore size. Highest activity of cell-free ice nuclei was obtained when cells were grown in low salinity (0.5-5% NaCl, w/v). Freezing tempera ture threshold, estimated to be below - 7 degreesC indicating class C nucle i, was not affected by medium salinity. Their density, as estimated by Perc oll density centrifugation, was 1.018 +/- 0.002 g ml(-1) and they were foun d to be free of lipids. Ice nuclei are released in the growth medium of rec ombinant H. elongata cells probably because of inefficient anchoring of the ice-nucleation protein aggregates in the outer membrane. The ice(+) recomb inant H. elongata cells could be useful for future use as a source of activ e cell-free ice nucleation protein.