154
V. Carginale et al.
Table 2. Amino acid composition of zinc-binding proteins from T bernacchii (MT) and
C. hamatus (Zn-bp)
Amino acid
Zn-bp
MT
C. hamatus (mol%)
T bernacchii
(mol %)
Cysa
4.75
30.1
Asp
10.92
6.8
Thr
5.43
11.2
Ser
5.98
14.3
Glu
17.57
4.3
Pro
8.00
5.7
Gly
11.99
11.9
Ala
6.94
2.1
Val
5.68
1.5
Met
1.63
0
lie
4.12
0
Leu
5.10
0
Tyr
1.98
0
Phe
2.05
0
His
1.94
0
Lys
3.88
12.1
Arg
1.89
0
aDetermined as cysteic acid
as far as concerns the zinc status: in fact, very little, if any, zinc-thionein
is present in the icefish liver, most of zinc being associated with a nonMT low-molecular weight protein. No information is available at the
moment on the role played by such a protein: apparently, it resembles a
family of still ill-characterized metal-binding proteins described in a
number of vertebrate and invertebrate species. The situation is entirely
different in the red-blooded fish where the predominant species is given
byMT.
Sequence and Structure of T. bernacchii MT
Fish MTs have a blocked amino terminus. Hence, in order to obtain
information on MT sequence and structure, MT cDNA was generated by
reverse-transcriptase polymerase-chain-reaction (RT-PCR) using as
primers oligo-dT and an oligonucleotide designed on the amino terminal
sequence of pis cine MT [22]. Electrophoresis of the PCR reaction showed
a band of about 350 bp. This fragment was eluted from the gel, ligated
into a pGEM-T plasmid and cloned in E. coli. The plasmid DNA obtained