ProteinDNA interactions

1
Protein-DNA interactions

• Non-Structure, non-Sequence Specific
• Z-DNA binding proteins
• Non-sequence specific interactions by sequence
  specific DNA binding proteins
• Sequence Specific

2
The Importance of non-specific DNA binding

• Specific binding is a competition between the
  target site and all other accessible DNA.
• Non-specific binding reduces the dimensionality
  of the search from a 3D volume to a 1D line.
  Speeds up finding target by as much as 1,000x.
• Sequence specific binding usually accompanied by
  conformational change.

3
Z DNA Binding proteins
Almost all contacts to DNA backbone.
Schwartz et al., NSB, 2001, 8, 9, p.761
4
Pox Viruses Contain a Z DNA binding protein
required for infectivity

• Yaba monkey tumor virus (YMTV) induces tumors in
  monkeys and humans.
• The viral protein E3L is oncogenic and
  antiapoptotic.
• It has 2 domains, one binds Z DNA and one binds
  dsRNA.

5
Common Features of Z DNA Binding Proteins
6
The lac repressor-DNA complex

• Protein dimer interacts with (approximately)
  palindromic DNA to repress transcription
• Binding of the inducer (lactose) relieves
  repression by reducing specificity for DNA

7
The lac repressor
Hinge
DBD
8
Overview of the specific lac repressor DNA complex
DBD generally poorly stable w/o DNA. The hinge is
unfolded in the absence of DNA. Hinge inserts
into minor groove causing an opening and bending
of DNA. Binding of inducer disrupts hinge-DNA
interaction.
9
How does sequence specific binding occur?
Hinge helix
Asn50
C-term helix
The dissociation of the lac repressor from DNA,
which presumably, is the inverse of the
association. First, the hinge helix unfolds,
followed by a H-bond between Asn50 and the DNA.
The loss of this H-bond destabilizes the
C-terminal helix which in turn destabilizes the
surrounding helices. Only the recognition helix
remains stable and this forms part of a
hydrophobic core of the protein. Val15 and a few
other hydrophobic residues stabilize this core.
If this core is disrupted, all interaction with
the DNA is lost and the protein dissociates. This
order was determined by measuring the exchange
rate of the backbone amide 1H.
10
DNA recognition by the Helix-Turn-Helix motif
This clearly means that there is a profound
reduction in the number of conformational states
from which exchange can occur. Kalodimos et al,
NSB, 2002, 9, p.193. Thus DNA binding in an
entropically unfavored process.
11
Sequence-specific protein DNA interactions.
Major groove
Minor groove
Hinge helix interaction.
12
Specific contacts (cont.)
13
The non-specific complex of lac repressor and DNA

Kalodimos et al, Science, 2004, 305, p. 386
14
Comparison of Protein-DNA Contacts
Specific
Non Specific
15
The DNA binding coordinate of the lac repressor
This coordinate follows perfectly the presumed
association pathway shown earlier.
16
Take-Home Lessons

• Sequence specific DNA binding is always in
  competition with non-sequence specific binding.
  The ratio is defined by the Boltzmann
  distribution and therefore is dependent on the
  energy difference between the 2 states.
• Non-sequence specific binding to DNA usually
  occurs via the backbone and involves week
  coulombic interactions. These type of
  interactions generally display a high level of
  dynamic behaviour and are therefore enthalpy
  driven.
• Sequence specific protein-DNA complexes are
  characterized by direct interaction between the
  bases and protein. These interactions may well be
  hydrophobic and coulombic. They are more rigid
  and usually involve conformational changes from
  the free state and since DS goes down, the
  enthalpic changes must be much larger than for
  non-specific interactions.