CRYOPROTECTIN, A CABBAGE PROTEIN PROTECTING THYLAKOIDS FROM
FREEZE-THAW DAMAGE
Silke Schilling (1), Hany A. M. Sror (2), Dirk K. Hincha
(3), Jürgen M. Schmitt (1), Carsten A. Köhn (1)
(1) Institut für Pflanzenphysiologie und Mikrobiologie,
Freie Universität, Königin-Luise-Str. 12-16, 14195
Berlin, Germany
(2) Permanent address: Biochemistry Department, Faculty of
Agriculture, Ain-shams University, Cairo,
Egypt
(3) Present address: Max-Planck Institut für Molekulare
Pflanzenphysiologie, 14424 Potsdam, Germany
We have purified a protein (cryoprotectin) from the leaves
of cold-acclimated savoy cabbage (Brassica
oleracea L. convar. capitata (L.) Alef. var. sabauda L.),
which protects thylakoids isolated from the leaves of
non-acclimated spinach (Spinacia oleracea) from freeze-thaw
damage. Sequencing of cryoprotectin revealed
the copurification of at least three isoforms. The sequence
data showed homology to the WAX9 protein
family, which belongs to the class of nonspecific lipid
transfer proteins (Hincha et al., Plant Physiology, Vol.
125, pp. 835-846). Antibodies raised against a synthetic
peptide derived from the WAX9 protein sequence
recognized a band of approximately 10 kDa in Western blots
of crude cryoprotectin preparations.
Cryoprotective activity could be precipitated from solution
by the antiserum. To clarify the function of the
different members of the gene family the five wax genes so
far known are cloned and expressed in E. coli.
Wax b expressed in E. coli had lipid transfer activity but
no cryoprotective activity. Wax e isolated from
cabbage cuticula also had lipid transfer activity but no
cryoprotective activity. In contrast highly purified
fractions of cryoprotectin showed no lipid transfer
activity but strong cryoprotective activity. Work is in
progress to clone further genes with the aim to identify
the cryoprotective member of this gene family.
presented at: 6TH INTERNATIONAL PLANT COLD HARDINESS
SEMINAR 30.6. - 5.7.2001, Helsinki, Finland