Chemistry Reference
In-Depth Information
Gln residues implicated in DNA recognition. The 5
sites were cleaved
strongly by all metal-peptide complexes. However, in contrast to 434R, they showed
a slight 5
′
- ACAA - 3
′
preference. Single-site mutants at Q
7
or Q
3
did
not affect the relative specifi city of the metal-peptide complexes, but changed the
affi nity. Two issues were considered to contribute in the reduction in specifi city. The
fi rst issue involved the intrinsic site selectivity of the metallointercalator resulting
in preference also for the sites 5
′
- ACGA - 3
′
>
5
′
- ACAA - 3
′
. The second issue addressed
the conformational fl exibility of appended peptides resulting in decrease of the
thermodynamic advantage to be gained from the ensemble of contacts available in
the DNA-binding protein or even promote nonspecifi c interactions with the phos-
phate backbone, so that the major contribution to DNA site specifi city would be
derived instead from the metal complex.
48
Another interesting study involved a peptide (7p) derived from the recognition
loop of the restriction endonuclease
MunI
from
Mycoplasma unidentifi ed
, which
was appended to the groove-binding complex [Ru(bpy)
2
(m - bpy)]
2+
(m - bpy = 4 -
methyl - 4
′
- ACA - 3
′
or 5
′
- ATG - 3
′
- bipyridine).
49
MunI
recognizes the palindromic hexanu-
cleotide sequence C/AATTG and cleaves as indicated by the slash. All amino acid
residues involved in sequence-specifi c interactions lie within a single short region
(Arg
115
- Gly
116
- Asn
117
- Ala
118
- His
119
- Glu
120
- Arg
121
= 7p).
50
A
1
H NMR study of the
interactions of the diastereomeric complexes L - and D - [Ru(bpy)
2
(m - bpy - 7p)]
2+
with
the oligonucleotide sequences d(5
′
- carboxylate - 2,2
′
′
- CGCGATCGCG - 3
′
)
2
and
d(5
′
- GCGCT-
TAAGCGC - 3
)
2
indicated binding of both isomers to the ends of both sequences.
Furthermore, the L-isomer exhibited additional NOE contacts to the major groove
protons of the GCT/CGA sequence of the decanucleotide and the central TT/AA
part of the dodecanucleotide. No sequence-specifi c contacts were indicated by the
experimental results, albeit intermolecular NOEs showed that the peptide was ori-
ented towards the DNA major groove. Thus, the appended peptide did not repro-
duce the recognition characteristics of the endonuclease that it was derived from.
This is another case in which the conformational fl exibility of the peptide moiety
did not allow formation of sequence-specifi c contacts, whereas in the case of d(5
′
′
-
GCGCTTAAGCGC - 3
′
)
2
the resultant conformers were bound to DNA via many
different orientations.
49
12.2.6
Summary
Tethering of a peptide to a transition metal complex, especially to a metallointer-
calator, increases the overall DNA-binding affi nity. Many studies have shown that
minimalist peptide domains are insuffi cient for tight binding and dimerization has
been utilized in order to enhance affi nity.
7
N.Y. Sardesai
et al.
have demonstrated
that a 100-fold excess of free peptide was incapable of competing with the corre-
sponding rhodium metallointercalator-peptide conjugate in binding to the DNA
target.
46
On the other hand, the peptide contributes substantially to the DNA-
binding affi nity, resulting in tighter binding compared to the metal complex.
48
Simple coordination metal complexes often lack hydrogen-bonding donors
or acceptors, thus undergoing their site-specifi c reactions solely on the basis of
shapes and symmetries.
51
Thus, in most cases, the appended peptide contributed
