A natural variant of type I antifreeze protein with four ice‐binding repeats is a particularly potent antifreeze

Chao, Heman; Hodges, Robert S.; Kay, Cyril M.; Gauthier, Sherry Y.; Davies, Peter L.

doi:10.1002/pro.5560050617

Cited by 67 publications

(71 citation statements)

References 27 publications

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“…We investigated the effect of increasing temperature on the secondary structure of the protein using CD. A sample was warmed slowly from 4 to 18°C, and several spectra were collected and averaged at each temperature (4,11,15, and 18°C). The first appreciable changes in the spectrum occurred at 18°C.…”

Section: Methodsmentioning

confidence: 99%

“…The type I AFP isoforms that are comprised of three 11-amino acid repeats (HPLC 6 and 8) produce a maximum TH activity of less than 1 Celsius degree at concentrations above 20 mg/ml, and are present in the flounder plasma at a concentration of ϳ10 -15 mg/ml, which produces a TH of ϳ0.7 Celsius degree (1). The four-repeat isoform (AFP-9) has higher specific TH activity, but is present at a much lower concentration, and therefore makes only a moderate contribution to the TH activity of the plasma (11). The colligative freezing point depression produced by plasma solutes and the TH produced by AFPs are directly additive.…”

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confidence: 99%

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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: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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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“…Many of the IBPs with known structures have been determined experimentally to have sequences that are composed of multiple short repeats (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19) residues in length) that in turn form repeating structural motifs. These structural motifs include a-helices and polyproline type II helices, but the majority of repetitive IBPs form a b-solenoid structure.…”

Section: Introduction To Repetitive Protein Structuresmentioning

confidence: 99%

“…1(A)]. 18 Spruce budworm AFP (SbwAFP) has a well-characterized isoform (501) that is 30 residues longer than most, with the extra sequence making exactly two 15-residue repeats that each makes a coil of the b-solenoid structure. 19 While detection of repeats is often not especially difficult, finding the correct "frame" of the repeats is important for establishing the repeating structure, or coil.…”

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Modeling repetitive, non‐globular proteins

et al. 2016

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While ab initio modeling of protein structures is not routine, certain types of proteins are more straightforward to model than others. Proteins with short repetitive sequences typically exhibit repetitive structures. These repetitive sequences can be more amenable to modeling if some information is known about the predominant secondary structure or other key features of the protein sequence. We have successfully built models of a number of repetitive structures with novel folds using knowledge of the consensus sequence within the sequence repeat and an understanding of the likely secondary structures that these may adopt. Our methods for achieving this success are reviewed here.

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“…34 RmAFP1 has only one disulfide bridge and the C. perlata AFP has none. 14 Furthermore, a type I fish AFP, with an ahelical structure and no cysteines, have been reported to have a T m of only 18 C. 35 Two different "strategies" for obtaining heat tolerant insect AFPs have been observed-high melting temperatures or great refolding abilities. As illustrated in Eq.…”

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Low thermodynamic but high kinetic stability of an antifreeze protein from Rhagium mordax

et al. 2014

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The equilibrium heat stability and the kinetic heat tolerance of a recombinant antifreeze protein (AFP) from the beetle Rhagium mordax (RmAFP1) are studied through differential scanning calorimetry and circular dichroism spectroscopy. In contrast to other insect AFPs studied with this respect, the RmAFP1 has only one disulfide bridge. The melting temperature, T m , of the protein is determined to be 28.5 C (pH 7.4), which is much lower than most of those reported for AFPs or globular proteins in general. Despite its low melting temperature, both biophysical and activity measurements show that the protein almost completely refolds into the native state after repeated exposure of 70 C. RmAFP1 thus appears to be kinetically stable even far above its melting temperature. Thermodynamically, the insect AFPs seem to be dividable in three groups, relating to their content of disulfide bridges and widths of the ice binding motifs; high melting temperature AFPs (high disulfide content, TxT motifs), low melting temperature but high refolding capability AFPs (one disulfide bridge, TxTxTxT motifs) and irreversibly unfolded AFPs at low temperatures (no disulfide bridges, TxTxTxTxT motifs). The property of being able to cope with high temperature exposures may appear peculiar for proteins which strictly have their effect at subzero temperatures. Different aspects of this are discussed.

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A natural variant of type I antifreeze protein with four ice‐binding repeats is a particularly potent antifreeze

Cited by 67 publications

References 27 publications

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

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

Modeling repetitive, non‐globular proteins

Low thermodynamic but high kinetic stability of an antifreeze protein from Rhagium mordax

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