Antifreeze protein pseudogenes

Davies, Peter L.; Gauthier, Sherry Y.

doi:10.1016/0378-1119(92)90373-w

Cited by 16 publications

(25 citation statements)

References 19 publications

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“…This sequence is similar to that at the N terminus of the predicted 5a gene product (IDPAAKAAAAA) (21), but differs in the presence of an additional residue and the conservative replacement of the basic residue. Amino acid analysis of the sample that had been purified to homogeneity using DEAE-cellulose revealed an exceptionally alanine-rich composition (Ͼ60 mol %) that was also abundant in threonine (13 mol %) but contained much lower quantities of the other amino acids, including leucine and phenylalanine that are typically associated with a hydrophobic core.…”

Section: Purification Of the 17-kda Afp-mentioning

confidence: 78%

Hyperactive Antifreeze Protein from Winter Flounder Is a Very Long Rod-like Dimer of α-Helices

Marshall

Chakrabartty

Davies

2005

Journal of Biological Chemistry

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The winter flounder (Pseudopleuronectes americanus) produces short, monomeric ␣-helical antifreeze proteins (type I AFP), which adsorb to and inhibit the growth of ice crystals. These proteins alone are not sufficiently active to protect this fish against freezing at ؊1.9°C, the freezing point of seawater. We have recently isolated a hyperactive antifreeze protein from the plasma of the flounder with activity 10 -100-fold higher than type I AFP. It is comparable in activity to the AFPs produced by insects, and is capable of conferring freeze resistance to the flounder. This novel AFP has a molecular mass of 16,683 Da and a remarkable amino acid composition that is >60% alanine. CD spectra indicate that the protein is almost entirely ␣-helical at 4°C but partially denatures at 20°C, resulting in a species with a moderately reduced helix content that is stable at up to 50°C. This transformation correlates with irreversible loss of activity. Analytical ultracentrifugation (sedimentation velocity and equilibrium) indicates that the predominant species in solution is dimeric (molecular weight, 32,275). Size-exclusion chromatography reveals a 2-fold higher apparent molecular weight suggesting that this molecule has an unusually large Stokes radius. The axial ratio of the dimer calculated from the sedimentation velocity data is 18:1, confirming that this protein has an extraordinarily long, rod-like structure, consistent with a novel dimeric ␣-helical arrangement. The structural model that best fits these data is one in which the ϳ195 amino acids of each monomer form one ϳ290-Å long ␣-helix and associate via a unique dimerization motif that is distinct from that of the leucine zipper and any other coiled-coil.The winter flounder is adapted to the harsh environment of the inshore North Atlantic, where it can live in ice-laden water at temperatures that potentially go as low as Ϫ1.9°C, the freezing point of seawater. The flounder body fluids, which have an equilibrium freezing point of only Ϫ0.8°C (1), resist freezing, and this has been attributed to the presence of antifreeze proteins (AFPs) 1 (reviewed in Ref. 2). An AFP (later named type I, Ref.3) was discovered in winter flounder 30 years ago (4) and has been extensively studied. Type I AFPs are small alanine-rich proteins of 37-48 residues that form a single amphipathic ␣-helix (5), with an underlying 11-amino acid repeat (6). The 11-amino acid periodicity within the ␣-helix forms an alanine-rich face with intimate surface/surface complementarity to a pyramidal plane {20 -21} of the ice lattice, which mediates ice binding (7,8). Like other AFPs, type I AFP is thought to prevent ice growth by an adsorption-inhibition mechanism (9): when AFPs are bound to the surface of an ice crystal, the addition of water molecules to the ice lattice is restricted to the exposed surfaces between protein molecules. This forces the ice to grow in less thermodynamically favored curved surfaces, thus lowering the non-equilibrium freezing point. Interaction of the AFP with the {20 -21} a...

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Section: Purification Of the 17-kda Afp-mentioning

confidence: 78%

Hyperactive Antifreeze Protein from Winter Flounder Is a Very Long Rod-like Dimer of α-Helices

Marshall

Chakrabartty

Davies

2005

Journal of Biological Chemistry

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“…This band was blotted onto a poly(vinylidene difluoride) membrane, excised, and sequenced by N‐terminal Edman degradation (Table 1). The 12 residues determined were clearly homologous to, but distinct from, the start of the winter flounder 5a‐like 17 kDa AFP and the theoretical gene product (5a) after which it was named [16]. For example, the seventh residue in ApAFP is Lys.…”

Section: Resultsmentioning

confidence: 99%

“…Although the new AFP is 10–100‐times more active in TH than the small type I AFPs, they are quite likely related as paralogs. The original 5a ‘pseudogene’ sequence, which is clearly homologous (based on its predicted N‐terminal sequence) to the large hyperactive AFPs of winter flounder, American plaice and yellowtail flounder was isolated from a winter flounder genomic library by hybridization to a type I AFP cDNA probe [16,24]. Moreover, the 5a and type I AFP genes share substantial sequence identity in the 5′ and 3′ untranslated regions of their genes.…”

Section: Discussionmentioning

confidence: 99%

Hyperactive antifreeze protein in flounder species

et al. 2005

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Antifreeze proteins (AFPs) are functionally defined by their ability to bind to the surface of ice and inhibit its growth, which causes a lowering of the freezing temperature below the equilibrium freezing ⁄ melting point [1]. This thermal hysteresis (TH) effect of AFPs enables teleost fishes to live in ice-laden polar and subpolar oceans where temperatures can reach the freezing point of the seawater ()1.9°C), which is over 1 C°colder than the freezing temperature of their hypotonic body fluids ()0.7 to )0.9°C). Without the protection of AFPs, fish are effectively supercooled in these waters and will freeze on contact with ice [2,3]. Thus, the recent acquisition of AFPs during the late stages of the teleost radiation has enabled some species to survive in and ⁄ or expand into the relatively new niche created by sea-level glaciation 1-20 million years ago [4]. Type I AFPs are small, monomeric, alanine-rich single a-helices that have an 11-amino-acid periodicity. They are one of five distinct nonhomologous types of AFP found in fishes [5] and they are present in some righteye flounders, including the winter flounder (Pseudopleuronectes americanus), yellowtail flounder (Limanda ferruginea) and Alaskan plaice (Pleuronectes quadritaberulatus) [6][7][8]. The AFP isoforms in the winter flounder have been particularly well characterized. One of them, the 37-amino-acid HPLC-6, was the first AFP to have its structure solved [9,10] and to have the ice plane to which it binds defined by ice-etching [8]. The differences between isoforms mainly lie in their length (the number of 11-amino-acid repeats being either three or four) and in the amino acid replacements on the less well conserved hydrophilic side of

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“…Antifreeze proteins (AFPs), 2 also known in the insect literature as thermal hysteresis proteins (THPs), are a family of proteins which have the ability to inhibit ice growth by binding to the ice surface in an adsorptioninhibition mechanism (1)(2)(3). This inhibition causes a depression of the noncolligative freezing point of a solution below its melting point, which is thermal hysteresis (TH).…”

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confidence: 99%