Fuerzas que participan en las

interacciones DNA-Proteína

Jorge Arevalo
2022
 Protein-DNA interactions

usually involve some degree of sequence specificity

Figure 8.21 The TATA-binding protein (TBP) has been cocrystallized with DNA.
Figure reproduced with permission from Voet, D., Voet, J., and Pratt, C. W. Fundamentals of Biochemistry. New York: Wiley, 1999. © (1999) John
Wiley & Sons, Inc.
Textbook of Biochemistry with Clinical Correlations, 7e edited by Thomas M. Devlin © 2011 John Wiley & Sons, Inc.
Structural information is now available on over 400 distinct
DNA-protein complexes, from a wide range of eukaryotic and
prokaryotic sources. Studies of the proteins themselves rarely
provide sufficient insight into the processes of recognition.
The determination of the human genome sequence in 2001 has
enabled reliable estimates to be made for the numbers of genes
with particular functions. Of the 30000 in total, 13.5 per cent (2308)
are proposed to be
involved in nucleic acid
binding, of which 6
per cent (1850) are
estimated to be
transcription factors.
 Non-specific binding
Electrostatic forces are long range and not
very specific. They rule the attraction between
the positively charges protein surface
(all DNA-binding domains have exposed
basic side chains) and the negatively charged
DNA phosphate backbone. Once protein and
DNA are nearby due to electrostatic
interactions, the other forces,
which are shorter-range, become effective.
These, and predominantly
hydrogen bonds between amino acid side
chains and nucleic acid bases,
determine the protein-DNA binding specificity.
Once protein
and DNA are
nearby due
to electrostatic
interactions,
the other forces,
which are
shorter-range,
become effective.
These, and
predominantly
hydrogen bonds
between amino
acid side
chains and
nucleic acid bases,
determine
the protein-DNA
binding specificity.
 Specific binding
(universal amino acid-base interactions)
The only regions where the bases are available for interaction are at
the floor of the grooves. These are paved with nitrogen and oxygen atoms
that can make hydrogen bonds with the side chains of a protein.
 1. Hydrogen bonds
• Possible binding sides of DNA base pairs
Donadores ( +) y Aceptores
de puentes de hidrógeno( −).
Existen posiciones que no
forman puentes de hidrógeno
pero pueden participar de
interacciones electrostáticas (
)
The B-DNA major groove is the richer of the two groove of the duplex DNA,
both in information content per se, and in its ability to facilitate discrimination
between DNA sequences, which is essential if the appropriate genes are
to be transcribed. Thus, the major groove is generally the site of direct
information readout. Nonetheless, the minor groove is an important target
for some regulatory and structural proteins, especially those that able
to deform DNA so that the minor groove becomes greatly expanded.
For GC pair, the major groove exposes a hydrogen-bond acceptor, G N7,
another acceptor, G O6, a hydrogen-bond donor, C NH4, and finally,
a hydrogen atom at C5. The minor groove displays a hydrogen-bond
acceptor G N3, a donor G NH2 and an acceptor C O2.
For the AT pair, the major groove gives the following sequence : an
acceptor A N7, donor A NH6, acceptor T O4, and a methyl group at T5.
The minor groove displays an acceptor, a hydrogen atom and a donor.
These patterns for potential hydrogen bonds are clearly quite different for
the different base pairs in the major groove, and they could easily be
recognized and distinguished by a protein molecule.
Clearly, the major groove is a much better candidate for
sequence-specific recognition than the minor groove for two reasons.
First, the major groove is wider than the minor, and the bases are thus
more accessible to a protein molecule. Second, the pattern of possible
hydrogen bonds from the edges of the base pairs to a protein are
more specific and discriminatory in the major groove than in the minor.

Only a rather limited number of base pairs is needed to provide

unique and discriminatory recognition sites in the major groove.
The above figure gives the color codes for the hexanucleotide recognition
sites of three different restriction enzymes - Eco RI, Bal I and Sma I. It is
clear that these patterns are quite different, and each can be uniquely
recognized by specific protein-DNA interactions.
Puentes de hidrógeno de Arg, Gln y Aspn
There is no general 1:1 amino acid : DNA base correspondence, and
recognition can sometimes occur in a wide variety.

Here the distribution of amino acid-base interactions in 129 protein-DNA

Structures (Luscombe et., 2001) :
Gua Cyt Ade Thy #sum
-----------------------------------------------------------------------------------
Arginine (R) 98 8 19 24 149
Lysine (K) 30 6 4 9 49
Serine (S) 12 2 1 3 18
Asparagine (N) 7 10 18 7 42
Glutamine (Q) 6 2 16 2 26
Glutamate (E) 1 10 1 0 12

#sum 154 38 59 45 296

The majority of interactions involve O6 and/or N7 atoms of guanine bases
forming hydrogen bonds with the charged ends of long flexible side chains
from the basic residues arginine or lysine, the amide residues glutamine and
Asparagine or the hydroxyl group of a serine.
Arg – Gua : a perfect H-bonding association
(33% of the total of amino acid-base pair interactions)

DNA-binding domain
of Tc3 transposase
from C elegans
residue : Arg C236

2 H-bond
acceptors 1.82 Å
1.96 Å

PDBcode: 1tc3
2 H-bond R = 2.45 Å
donors
R-factor = 0.234
guanidinium
moeity
 Asn/Gln – Ade : another frequent H-bonding association
(11% of the total of amino acid-base pair interactions)

Pit-1 Pou domain

residue : Asn A44
PDBcode : 1au7
R = 2.30 Å
R-factor = 0.230

one H-bond
donors

one H-bond
acceptors

one H-bond
acceptors
1.96 Å

2.13 Å

one H-bond
donors formamide
group
 3

Recently, Cheng et al. (2003) 2

6

have calculated all

geometrically
plausible H-bonding 2 5

arrangements between amino

2
acid and nucleic acid base
(DNA and RNA recognition). 1

They have found 32 possible

interactions, with 17
5
of which have been observed
in complex structures. 1
84 18
(The number of observed
Cases are indicated
here in red). 10 3

25
26 183 7
 Cation-p/H-bond stair motif involve two nucleobases and an amino acid side chain.
Its encompass three different types of interactions :
p-p stacking, H-bond and cation-p interactions.

DNA-binding domain
of Tc3 transposase PDBcode: 1tc3
from C elegans R = 2.45 Å
residues : Arg C236-A7-A8 R-factor = 0.234
Zinc finger protein Homeodomain
PDBcode : 1mey From drosophila
R = 2.20 Å PDBcode : 1fjl
R-factor = 0.224 R = 2.00 Å
R-factor = 0.198

Sap-1 ets domain

PDBcode : 1bc8
R = 1.93 Å
R-factor = 0.220

Methyltransferase
PDBcode : 6mht
R = 2.05 Å
R-factor = 0.186
• Protein can bind the DNA through the base, sugar, and
the phosphate group
• Hydrogen bonds with phosphate are not specific, but
with great importance in stabilizing the protein-DNA
complexes
• Guanine exposes the greatest number of potential
hydrogen-bonding atoms on the base edge(4 positions)
• The polar and charged residues of amino acids play a
central role
Arg > Lys > Ser > Thr; Asn and Gln
• Acidic residues are used sparingly Asp and Glu
• Only Gly makes a significant number of interaction
• Few interactions are produced by hydrophobic residues
• Favored amino acid-base hydrogen
bonds
Arg and Lys --- G, Asp and Glu --- A, Ser and His --- G
80% of Ser and Thr’s interactions are with the DNA
backbone
• Hydrogen bond geometries
Single 36.9%
Bidentate 33.8% ( two or more hydrogen bonds are
made with a base or base pair)
Complex 34.1% ( a protein residue binds more than
one step simultaneously)
Example: Bidentate interaction with Arg
 2. Van der waals contacts
• Comprise 64.9% of all protein-DNA
interactions
• Interactions with the DNA backbone (
sugar and phosphate) are most prominent
• Interactions with the phosphate group
dominate due to their high exposure on
the DNA surface
• T>A>G>C
• Arg, Thr, Phe, Ile, His, Cys
• Phe and His may have ring stacking
interactions with the base ring
• Cys in coordinating proteins has a high
propensity to contact the DNA backbone
• Glu, Ala, Leu, and Asp are less favored:
Glu and Asp: electrostatic interactions with
DNA
Ala and Leu: shortness of their side chains
 3. Water mediated bonds
• Nearly as common as direct hydrogen bonds
• 14.9% of all protein-DNA interactions
• 70% are with the DNA backbone, mostly
phosphate group
• Interactions with purine are common than with
pyrimidine
• Polar and charged amino acids are frequently
used: Arg, Lys, Asp, Glu, Ser and Thr
 3. Water mediated bonds
• Nearly as common as direct
hydrogen bonds
• 14.9% of all protein-DNA
interactions
• 70% are with the DNA
backbone, mostly phosphate
group
• Interactions with purine are
common than with pyrimidine
• Polar and charged amino acids
are frequently used: Arg, Lys,
Asp, Glu, Ser and Thr
 Summery
Amino acids Mode of interaction Recognized base
Hydrogen bond
Arg, Lys Multiple-donor G/complex
His Multiple-donor (bifurcate) G
Ser Multiple-donor (bifurcate) G
Acceptor + donor Complex
Asn Gln Acceptor + donor A/complex
Asp, Glu Multiple-acceptor Complex
Van der waals contacts
Phe, Pro Ring-stacking A, T
Thr Methyl contact T
Gly, Ala, Val, Leu, Iso, Tyr Many (nonspecific)

No Base contact
Cys, Met, Trp
Ejemplos de contactos DNA Proteína
Los contactos pueden ser por
dos ó una cara del DNA
 Represor lac

Factores con
Homeodominios

Figure 8.24 Helix-turn-helix proteins use one helix to bind in the major groove while the other supports that
binding through hydrophobic interaction.
Redrawn from Alberts, B., Bray, D., Lewis, J., Raff, M., Roberts, K., and Watson, J. Molecular Biology of the Cell. New York: Garland, 1994.
Textbook of Biochemistry with Clinical Correlations, 7e edited by Thomas M. Devlin © 2011 John Wiley & Sons, Inc.
Los dedos de Zn se estabilizan por el metal
 TFIIIA, SP1, Gal4, Superfamilia de receptores hormonales
esteroideos

Figure 8.25 Two different Zn finger motifs are found in transcription factors.
(a) Reproduced with permission from Voet, D., and Voet, J. G. Biochemistry, 2d ed. New York: Wiley, 1995. © (1995) John Wiley & Sons, Inc. Part
(b) and (c) generously supplied by C. Pabo, M.I.T.
Textbook of Biochemistry with Clinical Correlations, 7e edited by Thomas M. Devlin © 2011 John Wiley & Sons, Inc.
 Fos

Jun

CREB

Figure 8.26 Leucine zipper proteins bind to DNA as dimers.

Modified from Alberts, B., Bray, D., Lewis, J., Raff, M., Roberts, K., and Watson, J. Molecular Biology of the Cell. New York: Garland, 1994.

Textbook of Biochemistry with Clinical Correlations, 7e edited by Thomas M. Devlin © 2011 John Wiley & Sons, Inc.
Figure 8.27 Transcription factor dimer formation is mediated through helix loop helix interactions.
Modified from Alberts, B., Bray, D., Lewis, J., Raff, M., Roberts, K., and Watson, J. Molecular Biology of the Cell. New York: Garland, 1994.
Textbook of Biochemistry with Clinical Correlations, 7e edited by Thomas M. Devlin © 2011 John Wiley & Sons, Inc.
Ejemplo de unión no específica:
estructura del Nucleosoma
 Reference

• N.M. Luscombe et al, Nucl. Acid Res 29,

2860-2874 (2001).
• Luger et al, Nature 389, 251-260 (1997)
Evidencia experimental de unión de
proteínas al DNA
Footprinting y EMSA