Tài liệu Báo cáo khoa học: Hyperactive antifreeze protein in flounder species The sole freeze

Hyperactive antifreeze protein in flounder species

The sole freeze protectant in American plaice

Sherry Y. Gauthier1

, Christopher B. Marshall1

, Garth L. Fletcher3 and Peter L. Davies1,2

1 Department of Biochemistry, Queen’s University, Kingston, ON, Canada

2 Protein Function Discovery Group, Queen’s University, Kingston, ON, Canada

3 Ocean Sciences Centre, Memorial University of Newfoundland, St. John’s, NF, Canada

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 tem￾perature 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) [6–8]. The AFP isoforms in the win￾ter 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 replace￾ments on the less well conserved hydrophilic side of

Keywords

alpha-helix; antifreeze protein; freezing point

depression; ice

Correspondence

P. L. Davies, Department of Biochemistry,

and Protein Function Discovery Group,

Queen’s University, Kingston, ON,

K7L 3N6, Canada

Fax: +1 613 5332497

Tel: +1 613 5332983

E-mail: [email protected]

(Received 24 March 2005, revised 8 July

2005, accepted 12 July 2005)

doi:10.1111/j.1742-4658.2005.04859.x

The recent discovery of a large hyperactive antifreeze protein in the blood

plasma of winter flounder has helped explain why this fish does not freeze

in icy seawater. The previously known, smaller and much less active type I

antifreeze proteins cannot by themselves protect the flounder down to the

freezing point of seawater. The relationship between the large and small

antifreezes has yet to be established, but they do share alanine-richness

(> 60%) and extensive a-helicity. Here we have examined two other right￾eye flounder species for the presence of the hyperactive antifreeze, which

may have escaped prior detection because of its lability. Such a protein is

indeed present in the yellowtail flounder judging by its size, amino acid

composition and N-terminal sequence, along with the previously character￾ized type I antifreeze proteins. An ortholog is also present in American

plaice based on the above criteria and its high specific antifreeze activity.

This protein was purified and shown to be almost fully a-helical, highly

asymmetrical, and susceptible to denaturation at room temperature. It is

the only detectable antifreeze protein in the blood plasma of the American

plaice. Because this species appears to lack the smaller type I antifreeze

proteins, the latter may have evolved by descent from the larger antifreeze.

Abbreviations

AFP, antifreeze protein; ApAFP, American plaice antifreeze protein; IAP, ice affinity purification; TH, thermal hysteresis.

FEBS Journal 272 (2005) 4439–4449 ª 2005 FEBS 4439