ID IBP_LEUSY Reviewed; 261 AA. AC C7F6X3; DT 18-JUL-2018, integrated into UniProtKB/Swiss-Prot. DT 22-SEP-2009, sequence version 1. DT 24-JUL-2024, entry version 44. DE RecName: Full=Ice-binding protein {ECO:0000303|PubMed:20067781}; DE AltName: Full=Antifreeze protein {ECO:0000312|EMBL:ACU30806.1}; DE Short=AFP {ECO:0000305}; DE AltName: Full=LeIBP {ECO:0000303|PubMed:21795798}; DE Flags: Precursor; GN Name=AFP {ECO:0000312|EMBL:ACU30806.1}; OS Leucosporidium sp. (strain AY30) (Arctic yeast). OC Eukaryota; Fungi; Dikarya; Basidiomycota; Pucciniomycotina; OC Microbotryomycetes; Leucosporidiales; Leucosporidium. OX NCBI_TaxID=662878; RN [1] {ECO:0000312|EMBL:ACU30806.1} RP NUCLEOTIDE SEQUENCE [GENOMIC DNA]. RC STRAIN=AY30 {ECO:0000312|EMBL:ACU30806.1}; RA Lee J.K., Kang S.H., Kim H.J.; RT "Gene cloning of antifreeze protein (AFP) of Leucosporidium sp. AY30."; RL Submitted (JUN-2009) to the EMBL/GenBank/DDBJ databases. RN [2] RP NUCLEOTIDE SEQUENCE [MRNA], PROTEIN SEQUENCE OF 214-226, IDENTIFICATION BY RP MASS SPECTROMETRY, FUNCTION, SUBCELLULAR LOCATION, AND GLYCOSYLATION. RX PubMed=20067781; DOI=10.1016/j.cryobiol.2010.01.002; RA Lee J.K., Park K.S., Park S., Park H., Song Y.H., Kang S.H., Kim H.J.; RT "An extracellular ice-binding glycoprotein from an Arctic psychrophilic RT yeast."; RL Cryobiology 60:222-228(2010). RN [3] RP PROTEIN SEQUENCE OF 21-29, FUNCTION, SUBUNIT, SUBCELLULAR LOCATION, RP GLYCOSYLATION, SIGNAL SEQUENCE, AND CIRCULAR DICHROISM ANALYSIS. RX PubMed=22426061; DOI=10.1016/j.cryobiol.2012.02.014; RA Park K.S., Do H., Lee J.H., Park S.I., Kim E., Kim S.J., Kang S.H., RA Kim H.J.; RT "Characterization of the ice-binding protein from Arctic yeast RT Leucosporidium sp. AY30."; RL Cryobiology 64:286-296(2012). RN [4] RP PROTEIN SEQUENCE OF 21-25, FUNCTION, SUBCELLULAR LOCATION, GLYCOSYLATION, RP MASS SPECTROMETRY, AND CIRCULAR DICHROISM ANALYSIS. RX PubMed=23203635; DOI=10.1007/s00253-012-4594-y; RA Lee J.H., Lee S.G., Do H., Park J.C., Kim E., Choe Y.H., Han S.J., RA Kim H.J.; RT "Optimization of the pilot-scale production of an ice-binding protein by RT fed-batch culture of Pichia pastoris."; RL Appl. Microbiol. Biotechnol. 97:3383-3393(2013). RN [5] RP CRYSTALLIZATION. RX PubMed=21795798; DOI=10.1107/s1744309111018446; RA Park A.K., Park K.S., Kim H.J., Park H., Ahn I.Y., Chi Y.M., Moon J.H.; RT "Crystallization and preliminary X-ray crystallographic studies of the ice- RT binding protein from the Arctic [correction of Aantarctic] yeast RT Leucosporidium sp. AY30."; RL Acta Crystallogr. F 67:800-802(2011). RN [6] RP FUNCTION, AND BIOTECHNOLOGY. RX PubMed=22622645; DOI=10.1007/s12010-012-9739-z; RA Lee S.G., Koh H.Y., Lee J.H., Kang S.H., Kim H.J.; RT "Cryopreservative effects of the recombinant ice-binding protein from the RT arctic yeast Leucosporidium sp. on red blood cells."; RL Appl. Biochem. Biotechnol. 167:824-834(2012). RN [7] {ECO:0007744|PDB:3UYU, ECO:0007744|PDB:3UYV} RP X-RAY CRYSTALLOGRAPHY (1.57 ANGSTROMS) OF 21-261 OF NON-GLYCOSYLATED AND RP GLYCOSYLATED PROTEINS, FUNCTION, SUBUNIT, GLYCOSYLATION AT ASN-185, RP MUTAGENESIS OF SER-43; ALA-49; THR-65; THR-109; SER-147; SER-171; THR-198; RP THR-216; SER-222; ALA-234; THR-239 AND 244-PHE--LEU-261, AND CIRCULAR RP DICHROISM ANALYSIS. RC STRAIN=AY30 {ECO:0000303|PubMed:22303017}; RX PubMed=22303017; DOI=10.1074/jbc.m111.331835; RA Lee J.H., Park A.K., Do H., Park K.S., Moh S.H., Chi Y.M., Kim H.J.; RT "Structural basis for antifreeze activity of ice-binding protein from RT arctic yeast."; RL J. Biol. Chem. 287:11460-11468(2012). RN [8] RP X-RAY CRYSTALLOGRAPHY (1.34 ANGSTROMS) OF 21-53; 52-97 AND 100-243 OF RP CHIMERA WITH BACTERIAL ANTIFREEZE PROTEIN, FUNCTION, SUBUNIT, AND CIRCULAR RP DICHROISM ANALYSIS. RX PubMed=24699650; DOI=10.1107/s1399004714000996; RA Do H., Kim S.J., Kim H.J., Lee J.H.; RT "Structure-based characterization and antifreeze properties of a RT hyperactive ice-binding protein from the Antarctic bacterium Flavobacterium RT frigoris PS1."; RL Acta Crystallogr. D 70:1061-1073(2014). CC -!- FUNCTION: Confers freeze tolerance. Binds to the surface of ice CC crystals and inhibits their growth. Has low thermal hysteresis (TH) CC activity, which is the ability to lower the freezing point of an CC aqueous solution below its melting point (PubMed:20067781, CC PubMed:22303017, PubMed:22426061, PubMed:22622645, PubMed:23203635, CC PubMed:24699650). The TH activity of this protein is approximately 0.2 CC degrees Celsius at 50 uM and 0.3 degrees Celsius at 400 uM CC (PubMed:24699650). {ECO:0000269|PubMed:20067781, CC ECO:0000269|PubMed:22303017, ECO:0000269|PubMed:22426061, CC ECO:0000269|PubMed:22622645, ECO:0000269|PubMed:23203635, CC ECO:0000269|PubMed:24699650}. CC -!- SUBUNIT: Homodimer (PubMed:22303017, PubMed:22426061, PubMed:24699650). CC Dimerization is not required for the thermal hysteresis (TH) activity CC (PubMed:22303017). {ECO:0000269|PubMed:22303017, CC ECO:0000269|PubMed:22426061, ECO:0000269|PubMed:24699650}. CC -!- SUBCELLULAR LOCATION: Secreted {ECO:0000269|PubMed:20067781, CC ECO:0000269|PubMed:22426061, ECO:0000269|PubMed:23203635}. CC -!- PTM: Glycosylated (PubMed:20067781, PubMed:22303017, PubMed:22426061, CC PubMed:23203635). Glycosylation is not required for the thermal CC hysteresis (TH) activity (PubMed:22426061). Glycosylation may increase CC stability and secretion of this protein (Probable). CC {ECO:0000269|PubMed:20067781, ECO:0000269|PubMed:22303017, CC ECO:0000269|PubMed:22426061, ECO:0000269|PubMed:23203635, CC ECO:0000305|PubMed:22303017}. CC -!- MASS SPECTROMETRY: Mass=25565; Method=MALDI; Note=The measured mass is CC that of the mature protein.; Evidence={ECO:0000269|PubMed:23203635}; CC -!- BIOTECHNOLOGY: Improves cryopreservation of the human red blood cells CC (RBCs). Almost 90 % hemolysis damage induced to RBCs by freeze thawing CC with the slow warming at 22 degrees Celsius is dramatically decreased CC to less than 16 % when 0.4 or 0.8 mg/ml of this protein is used as a CC cryoprotectant. The post-thaw cell counts of RBCs frozen in the CC presence of 0.8 mg/ml of this protein and thawed by slow warming at 22 CC degrees Celsius is almost the same as that of non-frozen intact RBCs, CC whereas the recovery of RBCs frozen in its absence is less than 30 %. CC {ECO:0000269|PubMed:22622645}. CC -!- SIMILARITY: Belongs to the ice-binding protein family. {ECO:0000305}. CC -!- WEB RESOURCE: Name=Protein Spotlight; Note=Ice whisperer - Issue 209 of CC December 2018; CC URL="https://web.expasy.org/spotlight/back_issues/209/"; CC --------------------------------------------------------------------------- CC Copyrighted by the UniProt Consortium, see https://www.uniprot.org/terms CC Distributed under the Creative Commons Attribution (CC BY 4.0) License CC --------------------------------------------------------------------------- DR EMBL; GQ336994; ACU30806.1; -; Genomic_DNA. DR EMBL; GQ336995; ACU30807.1; -; mRNA. DR PDB; 3UYU; X-ray; 1.57 A; A/B=21-261. DR PDB; 3UYV; X-ray; 2.43 A; A=21-261. DR PDB; 4NU3; X-ray; 1.40 A; A/B=52-97. DR PDB; 4NUH; X-ray; 1.34 A; A=21-53, A=100-243. DR PDBsum; 3UYU; -. DR PDBsum; 3UYV; -. DR PDBsum; 4NU3; -. DR PDBsum; 4NUH; -. DR AlphaFoldDB; C7F6X3; -. DR SMR; C7F6X3; -. DR GlyCosmos; C7F6X3; 1 site, No reported glycans. DR iPTMnet; C7F6X3; -. DR EvolutionaryTrace; C7F6X3; -. DR GO; GO:0005576; C:extracellular region; IEA:UniProtKB-SubCell. DR GO; GO:0050825; F:ice binding; IDA:UniProtKB. DR InterPro; IPR021884; Ice-bd_prot. DR Pfam; PF11999; Ice_binding; 1. PE 1: Evidence at protein level; KW 3D-structure; Antifreeze protein; Direct protein sequencing; Glycoprotein; KW Secreted; Signal; Stress response. FT SIGNAL 1..20 FT /evidence="ECO:0000255, ECO:0000269|PubMed:22426061, FT ECO:0000269|PubMed:23203635" FT CHAIN 21..261 FT /note="Ice-binding protein" FT /evidence="ECO:0000255" FT /id="PRO_5011088933" FT SITE 65 FT /note="Ice-binding" FT /evidence="ECO:0000305|PubMed:22303017" FT SITE 147 FT /note="Ice-binding" FT /evidence="ECO:0000305|PubMed:22303017" FT SITE 234 FT /note="Ice-binding" FT /evidence="ECO:0000305|PubMed:22303017" FT CARBOHYD 185 FT /note="N-linked (GlcNAc...) asparagine" FT /evidence="ECO:0000269|PubMed:22303017, FT ECO:0007744|PDB:3UYV" FT MUTAGEN 43 FT /note="S->Y: Has 35% thermal hysteresis (TH) activity of FT that of the wild-type." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 49 FT /note="A->Y: Has 89% thermal hysteresis (TH) activity of FT that of the wild-type." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 65 FT /note="T->Y: Has 28% thermal hysteresis (TH) activity of FT that of the wild-type." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 109 FT /note="T->Y: Has 92% thermal hysteresis (TH) activity of FT that of the wild-type." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 147 FT /note="S->Y: Has 62% thermal hysteresis (TH) activity of FT that of the wild-type. Loss of TH activity; when associated FT with Y-234." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 171 FT /note="S->Y: Has 67% thermal hysteresis (TH) activity of FT that of the wild-type." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 198 FT /note="T->Y: Has 50% thermal hysteresis (TH) activity of FT that of the wild-type." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 216 FT /note="T->Y: Has 59% thermal hysteresis (TH) activity of FT that of the wild-type." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 222 FT /note="S->Y: Has 64% thermal hysteresis (TH) activity of FT that of the wild-type." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 234 FT /note="A->Y: Has 47% thermal hysteresis (TH) activity of FT that of the wild-type. Loss of TH activity; when associated FT with Y-147." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 239 FT /note="T->Y: No effect on thermal hysteresis (TH) FT activity." FT /evidence="ECO:0000269|PubMed:22303017" FT MUTAGEN 244..261 FT /note="Missing: Has 13% higher thermal hysteresis (TH) FT activity compared to the wild-type. Loss of FT homodimerization." FT /evidence="ECO:0000269|PubMed:22303017" FT HELIX 29..33 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 35..46 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 49..52 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 54..59 FT /evidence="ECO:0007829|PDB:3UYU" FT HELIX 61..63 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 64..67 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 83..85 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 87..89 FT /evidence="ECO:0007829|PDB:3UYU" FT HELIX 96..116 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 121..124 FT /evidence="ECO:0007829|PDB:3UYU" FT HELIX 125..128 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 132..135 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 137..144 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 146..148 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 152..155 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 162..168 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 170..172 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 177..181 FT /evidence="ECO:0007829|PDB:3UYU" FT HELIX 186..188 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 189..195 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 197..199 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 204..213 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 215..217 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 222..237 FT /evidence="ECO:0007829|PDB:3UYU" FT STRAND 239..241 FT /evidence="ECO:0007829|PDB:3UYU" SQ SEQUENCE 261 AA; 26807 MW; F114BC33CBC9B469 CRC64; MSLLSIITIG LAGLGGLVNG QRDLSVELGV ASNFAILAKA GISSVPDSAI LGDIGVSPAA ATYITGFGLT QDSSTTYATS PQVTGLIYAA DYSTPTPNYL AAAVANAETA YNQAAGFVDP DFLELGAGEL RDQTLVPGLY KWTSSVSVPT DLTFEGNGDA TWVFQIAGGL SLADGVAFTL AGGANSTNIA FQVGDDVTVG KGAHFEGVLL AKRFVTLQTG SSLNGRVLSQ TEVALQKATV NSPFVPAPEV VQKRSNARQW L //