Isolation and Characterization of Skin-type, Type I Antifreeze Polypeptides from the Longhorn Sculpin, Myoxocephalus octodecemspinosus *

The antifreeze polypeptides (AFPs) are found in several marine fish and have been grouped into four distinct biochemical classes (type I-IV). Recently, the new subclass of skin-type, type I AFPs that are produced intracellularly as mature polypeptides have been identified in the winter flounder (Pleuronectes americanus) and the shorthorn sculpin (Myoxocephalus scorpius). This study demonstrates the presence of skin-type AFPs in the longhorn sculpin (Myoxocephalus octodecemspinosus), which produces type IV serum AFPs. Using polymerase chain reaction-based methods, a clone that encoded for a type I AFP was identified. The clone lacked a signal sequence, indicating that the mature polypeptide is produced in the cytosol. A recombinant protein was produced in Escherichia coli and antifreeze activity was characterized. Four individual Ala-rich polypeptides with antifreeze activity were isolated from the skin tissue. One polypeptide was completely sequenced by tandem MS. This study provides the first evidence of a fish species that produces two different biochemical classes of antifreeze proteins (type I and type IV), and enforces the notion that skin-type AFPs are a widespread biological phenomenon in fish.

[1]  R. Hodges,et al.  New ice‐binding face for type I antifreeze protein , 1999, FEBS letters.

[2]  A. Haymet,et al.  Type I 'antifreeze' proteins. Structure-activity studies and mechanisms of ice growth inhibition. , 1999, European journal of biochemistry.

[3]  C. Hew,et al.  Secretory expression and site-directed mutagenesis studies of the winter flounder skin-type antifreeze polypeptides. , 1999, European journal of biochemistry.

[4]  C. Hew,et al.  Studies of a putative ice‐binding motif in winter flounder skin‐type anti‐freeze polypeptide , 1999, FEBS letters.

[5]  C. Cheng,et al.  Evolution of the diverse antifreeze proteins. , 1998, Current opinion in genetics & development.

[6]  R. Laursen,et al.  Isolation and characterization of an antifreeze protein from the longhorn sculpin, Myoxocephalus octodecimspinosis. , 1998, Biochimica et biophysica acta.

[7]  G. Fletcher,et al.  Skin-type Antifreeze Protein from the Shorthorn Sculpin,Myoxocephalus scorpius , 1998, The Journal of Biological Chemistry.

[8]  R. Laursen,et al.  Cloning and sequencing of cDNA encoding the LS-12 antifreeze protein in the longhorn sculpin, Myoxocephalus octodecimspinosis. , 1998, Biochimica et biophysica acta.

[9]  K. Merz,et al.  Ice-binding mechanism of winter flounder antifreeze proteins. , 1997, Biophysical journal.

[10]  D J Lipman,et al.  Making (anti)sense of non-coding sequence conservation. , 1997, Nucleic acids research.

[11]  R. Laursen,et al.  Amino acid sequence of a new type of antifreeze protein, from the longhorn sculpin Myoxocephalus octodecimspinosis , 1997, FEBS letters.

[12]  A. E. Oliver,et al.  Antifreeze glycoproteins inhibit leakage from liposomes during thermotropic phase transitions. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[13]  Y. Yeh,et al.  Antifreeze Proteins: Structures and Mechanisms of Function. , 1996, Chemical reviews.

[14]  C. Hew,et al.  Skin Antifreeze Protein Genes of the Winter Flounder, Pleuronectes americanus, Encode Distinct and Active Polypeptides without the Secretory Signal and Prosequences (*) , 1996, The Journal of Biological Chemistry.

[15]  F. Sicheri,et al.  Ice-binding structure and mechanism of an antifreeze protein from winter flounder , 1995, Nature.

[16]  L. Duret,et al.  Strong conservation of non-coding sequences during vertebrates evolution: potential involvement in post-transcriptional regulation of gene expression. , 1993, Nucleic acids research.

[17]  R. Laursen,et al.  Structure-function relationships in an antifreeze polypeptide. The role of neutral, polar amino acids. , 1992, The Journal of biological chemistry.

[18]  P. Davies,et al.  Antifreeze protein pseudogenes. , 1992, Gene.

[19]  C. Hew,et al.  Protein interaction with ice. , 1992, European journal of biochemistry.

[20]  A. Chakrabartty,et al.  The effect of enhanced α‐helicity on the activity of a winter flounder antifreeze polypeptide , 1991 .

[21]  C. Hew,et al.  Biochemistry of fish antifreeze proteins , 1990, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[22]  A. Chakrabartty,et al.  Structure-function relationship in a winter flounder antifreeze polypeptide. II. Alteration of the component growth rates of ice by synthetic antifreeze polypeptides. , 1989, The Journal of biological chemistry.

[23]  A. Chakrabartty,et al.  Crystal structure of an antifreeze polypeptide and its mechanistic implications , 1988, Nature.

[24]  A. Chakrabartty,et al.  Primary structures of the alanine-rich antifreeze polypeptides from grubby sculpin, Myoxocephalus aenaeus , 1988 .

[25]  T. Burcham,et al.  Antifreeze glycoproteins from polar fish blood. , 1986, Annual review of biophysics and biophysical chemistry.

[26]  V. Ananthanarayanan,et al.  Structures of shorthorn sculpin antifreeze polypeptides. , 1985, European journal of biochemistry.

[27]  A. Devries.  Antifreeze peptides and glycopeptides in cold-water fishes. , 1983, Annual review of physiology.

[28]  V. Ananthanarayanan,et al.  Antifreeze proteins from the shorthorn sculpin, Myoxocephalus scorpius: isolation and characterization. , 1980, Canadian journal of biochemistry.