Structure
of Cro and cI Repressors
Cro Structure
- Cro is a homodimer of 66-residue
subunits folded into three 
-helical regions and three
strands (Figure 26.28).
Two of the helices, numbered 2 and 3 in the figure, are separated
by a short turn, forming a helix-turn-helix motif
(see here and here).
In the Cro dimer the two number 3 helices are 3.4 nm apart,
the length of one turn of the DNA double helix. This distance
is consistent with the two subunits binding on the same side of
the DNA helix, in adjacent major groove sites, with the number
3 helices lying lengthwise in the grooves.
The amino acid sequence within and between helices 2 and 3 show remarkable homology with corresponding sequences in a large family of sequence-specific DNA-binding proteins but not with DNA-binding proteins that showed no sequence preference. The helix-turn-helix motif is a commonly evolved structural element in transcriptional regulatory proteins, at least in prokaryotes. A different motif, the zinc finger, is present in a large number of eukaryotic DNA-binding proteins (and a few known prokaryotic proteins). Other structural patterns of DNA-binding proteins, including the helix-loop-helix and the leucine zipper, have been described. These motifs are discussed here.
cI Structure
- The 
cI repressor also has
a helix-turn-helix motif. The relevant three-dimensional similarities
for Cro, cAMP receptor protein (CRP, see here),
and cI repressor are shown in Figure
26.29. cI repressor contains an additional binding
determinant-a pair of "arms," or short polypeptide segments
that extend from helix 1. They can be seen in Figure
26.1, extending around the helix and establishing contacts
on the other side of the DNA duplex. These arms probably explain
why cI binds more tightly to its operators than Cro
does.
The -3 helix of cI is
called the recognition helix, because when cI is bound
to DNA, its position deep within the major groove allows it to
contact specific DNA bases and hence to determine sequence specificity
of binding. The -2 helix is in contact primarily with
DNA phosphates. These electrostatic contacts strengthen binding
but do not contribute to specificity.
Amino acids and DNA binding - Amino acid sequence homologies among the DNA binding regions for Cro and cI are shown in Figure 26.30. The sequences, though similar, are not identical, which explains how Cro and cI repressors differ in their relative affinities for different operators. The residues common to both proteins are in contact with DNA sequence elements common to all of the operators (Figure 26.31). In both proteins a glutamine residue interacts with one A-T base pair, as shown. cI repressor, which binds most strongly to OR1, establishes specificity through a contact in OR1 with a unique alanine residue, whereas Cro, which binds most strongly to OR3, can be in contact with three specific base pairs in OR3 with unique asparagine and lysine residues.
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