title: Isolation and characterization of antifreeze proteins from the antarctic marine microalga Pyramimonas gelidicola.
abstract: Antifreeze proteins (AFPs) play an important role in the psychrophilic adaptation of polar organisms. AFPs encoded by an Antarctic chlorophyte, identified as Pyramimonas gelidicola, were isolated and characterized. Two AFP isoforms were found from cDNAs and their deduced molecular weights were estimated to be 26.4 kDa (Pg-1-AFP) and 27.1 kDa (Pg-2-AFP). Both AFP cDNAs were cloned and expressed in Escherichia coli. The purified recombinant Pg-1-rAFP and Pg-2-rAFP both showed antifreeze activity based on the measurement of thermal hysteresis (TH) and morphological changes to single ice crystals. Pg-1-rAFP shaped ice crystals into a snowflake pattern with a TH value of 0.6 ± 0.02 °C at ~15 mg/ml. Single ice crystals in Pg-2-rAFP showed a dendritic morphology with a TH value of 0.25 ± 0.02 °C at the same protein concentration. Based on in silico protein structure predictions, the three-dimensional structures of P. gelidicola AFPs match those of their homologs found in fungi and bacteria. They fold as a right-handed β-helix flanked by an α-helix. Unlike the hyperactive insect AFPs, the proposed ice-binding site on one of the flat β-helical surfaces is neither regular nor well-conserved. This might be a characteristic of AFPs used for freeze tolerance as opposed to freeze avoidance. A role for P. gelidicola AFPs in freeze tolerance is also consistent with their relatively low TH values.
authors.affiliation: Department of Life Science, Division of Natural Sciences, Hanyang University, 133-791, Seoul, South Korea.
authors.forename: Woongsic
authors.initials: W
authors.lastname: Jung
authors.forename: Yunho
authors.initials: Y
authors.lastname: Gwak
authors.forename: Peter L
authors.initials: PL
authors.lastname: Davies
authors.forename: Hak Jun
authors.initials: HJ
authors.lastname: Kim
authors.forename: EonSeon
authors.initials: E
authors.lastname: Jin
mesh_terms: D000595:Amino Acid Sequence; D000864:Antarctic Regions; D021301:Antifreeze Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D000460:Chlorophyta / Q000235:genetics* / Q000254:growth & development* / Q000378:metabolism; D003001:Cloning, Molecular; D016000:Cluster Analysis; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli; D005822:Genetic Vectors / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames / Q000235:genetics; D010802:Phylogeny; D017510:Protein Folding; D020033:Protein Isoforms / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA
chemical_list: D021301:Antifreeze Proteins; D017931:DNA Primers; D018076:DNA, Complementary; D020033:Protein Isoforms; D011994:Recombinant Proteins