334 Current Chemical Biology, 2009, 3, 334-342

NS3 Helicases as Drug Targets

Patrick Chène*

Disease Area Oncology, Novartis Institutes for BioMedical Research, CH-4002 Basel, Switzerland

Abstract: Members of the viral family Flaviviridae are a major cause of infectious diseases, such as hepatitis C, Dengue
fever, Japanese encephalitis, Yellow fever or West Nile fever. Major efforts are therefore being made to identify medi-
cines that can fight these infections. The genome of the Flaviviridae contains a single ORF that is translated into a precur-
sor polyprotein which is processed in the host into several proteins. Amongst these, the nonstructural protein 3 (NS3) is
formed from two domains that show distinct catalytic activities: a serine protease and a helicase activity. Since both ac-
tivities are required for viral replication, there is a lot of interest in investigating NS3 as a potential drug target. On the ba-
sis of the structural information available, we shall assess here the druggability of the helicase domain of NS3.

Keywords: NS3 helicase, hepatitis C virus, dengue virus, west nile virus.

1. INTRODUCTION these enzymes can be targeted by low molecular weight

The viral family Flaviviridae is divided into three gen-
era: Flavivirus, Hepacivirus and Pestivirus [1]. Amongst
Flaviviridae, the genus Flavivirus encompasses the largest
number of species. Flaviviridae are at the root of several
human diseases, such as Dengue fever, Yellow fever, Japa-
nese encephalitis and Hepatitis C [2, 3]. These diseases af-
fect a large number of individuals all over the world and
major efforts are being made to identify therapeutic agents
for these infections. One of the strategies which is currently
under investigation is the identification of low molecular
weight molecules that could selectively block the function of
the proteins encoded by these viruses.
The organization and length of the genome is similar for
all Flaviviridae. It comprises a ~9.5 to ~12.5 kb single-
stranded positive sense RNA that forms a single open read-
ing frame. The translation of the viral genome in the host cell
leads to the synthesis of a large single polyprotein. This pro-
tein is then processed by both viral and host proteases to
produce several viral proteins. These include three structural
proteins called C, prM and E, and seven nonstructural pro-
teins called NS1, NS2A, NS2B, NS3, NS4A, NS4B and
NS5. Several of these proteins possess a catalytic activity
and are therefore attractive targets for drug discovery. NS5
possesses S-adenosyl methyltransferase and RNA-dependent
polymerase activities, both being potentially interesting tar-
gets for molecules that inhibit these catalytic activities [4-7].
The second viral protein which is currently under investiga-
tion is the NS3 protein. This also possesses multiple catalytic
activities: serine protease, helicase and RNA triphosphatase.
Proteases are classical drug discovery targets, and substantial
efforts have been made to identify inhibitors of the
Flaviviridae proteases [7-10]. Helicases are rather new tar-
gets for drug discovery, and these enzymes have not yet been
intensively investigated by the pharmaceutical companies.
However, some recent examples nicely demonstrate that
*Address correspondence to this author at the Disease Area Oncology,
Novartis Institutes for BioMedical Research, CH-4002 Basel, Switzerland;
Tel: +41616962050; Fax: +41616963835;
E-mail: patrick_chene@yahoo.com
compounds [11, 12]. This opens up new opportunities for
drug discovery, since helicases form a large family of pro-
teins present in bacteria, viruses and humans.
Helicases catalyze the separation of double-stranded
DNA or RNA into single-strands in an ATP-dependent man-
ner [13]. For unwinding double-stranded nucleic acids, heli-
cases translocate along nucleic acids. Several mechanisms
have been proposed to explain the movement of helicases on
nucleic-acids [14-18]. It should be noted that helicases have
to be differentiated from translocases that just move onto
nucleic-acids without inducing strand separation. Helicases
possess several primary sequence stretches, called motifs,
that are well conserved and that have been used to classify
them. On the basis of these motifs and of structural consid-
erations, helicases are grouped into different superfamilies
[19]. The NS3 helicases belong to superfamily II. This is the
largest helicase superfamily, and enzymes belonging to su-
perfamily II are involved in broad variety of processes re-
lated to nucleic acid biology. Many of these proteins, includ-
ing NS3, are RNA helicases and, within the superfamily II,
they are classified into the DExD/H helicases family [20].
The name of this family comes from the primary sequence of
a motif called motif 2 or Walker B [21], which is conserved
in these enzymes. This small region is used by helicases to
coordinate the divalent cation (magnesium) used as co-factor
for ATP hydrolysis during the catalytic cycle [22]. The other
conserved motifs (in total 7) found in helicases are either
involved in the binding of the nucleotide (ATP) or in the
interaction with nucleic acids [19, 20, 22].
The fact that NS3 helicases are enzymes and that they are
important for the life cycle of Flaviviridae is a necessary but
not sufficient prerequisite to make them suitable targets for
drug discovery. A very important aspect of drug discovery is
the druggability of the selected target, which is a fuzzy no-
tion. It is linked to the possibility of identifying binding
site(s) on the target protein, where potent, low molecular
weight compounds could bind and affect its biological func-
tion with the desired outcome. The word potent is quite im-
portant, because it is potency which ensures that the selected
target is really druggable. The potency required in drug dis-

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NS3 Helicases as Drug Targets Current Chemical Biology, 2009, Vol. 3, No. 3 335

covery is defined by the efficacy of compounds in humans. It
is often possible to identify weakly active molecules that
affect the selected target, but if its druggability is low, the
optimization of these initial leads to produce more potent
compounds might be very difficult.
The best way to estimate the druggability of a protein is
to look at its three dimensional structure in order to see if
possible compound binding sites can be identified on its sur-
face. Until recently, few X-ray structures of NS3 helicases
were available, but in the last years several of them have
been published, and it is now possible to study these proteins
in more detail. In this short review, we shall describe the
general structure of NS3 helicases and evaluate different
possible strategies that could be used to target these en-
zymes.
2. GENERAL STRUCTURE OF THE NS3 HELICASE
Several structures of NS3 helicase are available (Table
1). These enzymes have been crystallized in their apo con-
formation or in the presence of a nucleotide and/or a nucleic
acid (NA). Structures of full-length NS3 proteins were also
determined (hereafter, NS3 corresponds to the full-length
NS3 protein, including both the protease and helicase do-
mains; NS3hel corresponds to the helicase domain alone).
Detailed descriptions of these structures are provided in the
original publications, and readers interested in obtaining a
deeper knowledge should consult these references (Table 1).
We shall focus here on the general description of NS3hel.
An alignment of the primary sequences of the different
NS3hel that have been crystallized is provided in Fig. (1).
No helicase structure from viruses belonging to the genus
Pestivirus is available. Nevertheless, the primary sequence
of the NS3hel from border disease virus was also included
for comparison. The primary sequence homology amongst
the different NS3hel from flaviviruses is quite high. There is
a lower degree of homology between these enzymes and the
HCV NS3hel (genus Hepacivirus) and an even lower one
with the BDV NS3hel (genus Pestivirus). Some of the motifs
(1, 2, 3 and 6) characteristic of helicases [20, 22] are well
conserved but a lower homology is observed between motifs
1A, 4 and 5.
The structures of DENV and HCV NS3hel, as prototype
helicases from viruses from their respective genera, are rep-
resented in Fig. (2). The helicase domain is formed of three
subdomains. Subdommmain 1 (white, Fig. 2) and subdomain
2 (grey, Fig. 2) have a structure similar to that of the RecA
protein [23]. These two subdomains are linked by a flexible
linker. It has been observed that the distance between sub-
domain 1 and subdomain 2 is different between the JEV and
HCV helicases [24]. Recently it has been shown that RNA
binding to the DENV helicase induces closure between
subdomains 1 and 2 [25]. The superimposition of different
structures (1hei and 8ohm) of the HCV helicase in its apo
conformation also shows that the distance between both sub

Table 1. Published X Ray Structures of NS3 Protein Constructs. For Simplicity the Serotypes for the Dengue and Hepatitis Vi-
ruses are not Indicated

Virus Protein form PDB code Resolution (Å) Reference

Flavivirus

Murray Valley encephalitis Apo 2v8o 1.9 [56]

Apo 1yks 1.8
Yellow fever [43]
ADP 1ymf 2.6
Apo 2bmf, 2bhr, 2jlq 2.4, 2.8, 1.7
ADPNP 2jlr 2
ADP 2jls 2.2
RNA 2jlu, 2jlw 2, 2.6
Dengue ADPNP; RNA 2jlv 1.9 [25, 27, 31]

ADP•VO4; RNA 2jlx 2.2

ADP•PO4 ; RNA 2jly 2.4
ADP; RNA 2jlz 2.2
Protease/Helicase 2vbc 3.15
Japanese encephalitis Apo 2z83 1.8 [24]
Kunjin virus Apo 2qeq 3.1 [57]
Kokobera virus Apo 2v6i 2.1 [58]
Hepacivirus
RNA 1a1v, 2f55 2.2, 3.3 [51, 59]
HCV Apo 1hei, 8ohm 2.1, 2.3 [60, 61]
Protease/Helicase 1cu1 2.5 [32]
336 Current Chemical Biology, 2009, Vol. 3, No. 3 Patrick Chène

Fig. (1). Alignment of the primary sequence of different NS3hel. Sequence start was chosen for all proteins on the basis of the location of the
linker region between the protease and helicase domain described for DENV NS3 [31]. MVEV: Murray Valley encephalitis virus; YFV: Yel-
low fever virus; DENV: Dengue virus; JEV: Japanese encephalitis virus; KUNV: Kunjin virus; KOKV: Kokobera virus; HCV: Hepatitis C
virus; BDV: Border disease virus. The position of the different helicase motifs is indicated on the basis of previous assignment established
from the primary sequence of HCV NS3 [20, 22]. The alignment was made using T-Coffee [62].
NS3 Helicases as Drug Targets Current Chemical Biology, 2009, Vol. 3, No. 3 337

domains varies for the same protein (data not shown). Alto-
gether this suggests that subdomains 1 and 2 can move rela-
tive to each other and that their movement may be linked to
different steps in the catalytic cycle [26]. Subdomain 3
(black, Fig. 2) is linked to subdomain 2 via a linker, and the
structure of subdomain 3 differs from that of the other two
subdomains. There is a lower homology between subdomain
3 from HCV and DENV NS3hel than between the two other
subdomains, which are more conserved. It has been postu-
lated that the lower similarity between subdomains 3 may
reflect different types of interaction between the NS3 protein
and one of its interacting partners, the NS5B protein [8, 27].
Several publications show that HCV NS3 also interacts via
its N-terminus with HCV NS5B [28] and that the protease
domain of NS3 is required for this interaction between these
two proteins [29, 30].
The structure the NS3 protein has been determined for
DENV and HCV [31, 32]. These structures are schematically
presented in Fig. (2C) and (2D). The position of the protease
domain (black on Fig. 2) is very different in the two proteins.
In the case of the DENV NS3 protein, the protease domain is
located at the opposite side of the C-terminus (sphere in Fig.
2C) of the NS3 protein, and the enzyme has an elongated
structure. In the case of the HCV NS3, both the protease and
the C-terminus of the NS3 protein are on the same side of the
enzyme, which has a more compact conformation (Fig. 2B).
This structural difference may have some important conse-
quences for the mechanism behind the processing of the viral
polyprotein by the NS3 protease [31, 33].
In addition to these observations, it has been shown that
the protease domain influences the catalytic activity of the
helicase domain. HCV NS3hel is unwinding RNA or DNA
more slowly than NS3 [30, 34, 35]. A similar observation
was made with DENV type 2 NS3 [36]. HCV NS3 hydro-
lyzes ATP more slowly than NS3hel when translocating on
nucleic acids, and NS3 assembles as higher oligomeric forms
than NS3hel on DNA and on ssDNA, suggesting a coopera-
tive effect in the presence of the protease domain [34]. Ex-
periments conducted with HCV NS3hel also indicate that
multiple molecules of this protein assemble on DNA [16,
37]. Beran et al. have proposed that the enhanced helicase
activity observed with NS3 when compared to NS3hel is a

Fig. (2). Structure of the NS3 protein. Solid ribbon representation of the DENV (A), HCV (B), DENV (C) and HCV (D) NS3hel proteins.
Within the helicase domain, subdomain 1, subdomain 2 and subdomain 3 are represented in white, grey and black, respectively (A, B). The
protease and the helicase domain are represented in black and white, respectively (C, D). The spheres represent the C-terminus of the en-
zymes. The structures used to make the figure are 2jlq (DENV NS3hel [25]), 1hei (HCV NS3 hel [61]), 2vbc (DENV NS3 [31]) and 1cu1
(HCV NS3 [32]).
338 Current Chemical Biology, 2009, Vol. 3, No. 3
consequence of a specific allosteric contribution from an
interaction between the helicase and protease domain [38].
Altogether these findings indicate that the catalytic activity
of the helicase domain of NS3 is influenced by the protease
domain. Therefore screens for inhibitors of NS3 helicase
activity should preferentially be done with NS3 and not with
the isolated helicase domain, NS3hel.
Other viral proteins seem to contribute to the regulation
of the catalytic activity of NS3. This is the case with NS4A
cofactor [39-41] and with NS5B [29, 30, 36]. NS2B also
modulates NS3hel activity [36, 42]. For example, it influ-
ences the selectivity of the WNV NS3 protein. WNV NS3
and WNV NS3hel unwind both DNA and RNA templates,
while WNV NS2B-NS3 only unwinds RNA templates and
has its DNA unwinding activity repressed [42]. The NS2B
co-factor enhances the affinity of DENV NS3 for ATP [31].
The structure of DENV fused to 18 residues of NS2B reveals
that a basic region located at the surface of the protease do-
main may increase the affinity for the nucleotide and partici-
pate in RNA binding [31]. Altogether these different results
indicate that NS3 helicase activity may be modulated by
various viral proteins and potentially by host proteins. On the
basis of our current information, it is difficult to predict
whether these different interactions can affect the binding of
compounds to NS3hel. Keeping this in mind, we shall now
discuss the different possible strategies for targeting NS3hel.
3. TARGETING THE NS3 HELICASE
Since NS3 helicases interact with both nucleotides and
NAs, two strategies can be proposed to inhibit these en-
zymes. The first is to identify compounds that prevent nu-
cleotide binding and/or hydrolysis. The second is to obtain
molecules that prevent NA binding and/or processing. In
both cases, the inhibitors could either directly compete with
the substrates for binding to NS3, or alternatively they could
interact with the protein on a remote site preventing by an
“allosteric” effect ATP and/or NA binding. Another possibil-
ity would be to identify compounds that, upon binding to the
NA, prevent its interaction with NS3hel. This approach, even
if it may lead to the identification of inhibitors, might be
Fig. (3). ATP binding site of DENV NS3hel. A. Surface representation of the DENV NS3hel protein. Subdomain 1, subdomain 2 and sub-
domain 3 are colored white, grey and black, respectively. The ATP analog, ADPNP, is represented at the interface between subdomain 1 and
subdomain 2. B. Details of the interaction between DENV NS3hel and the bound ADPNP. Water molecules are represented by small white
spheres and the metal cofactor (Me2+) by a grey sphere. For clarity, H bonds are not indicated. The structure used to make this representation
is 2jlv [25].
Patrick Chène
difficult to pursue, because it could be challenging to design
selective inhibitors, and therefore such molecules may show
some toxicity. This strategy will not be discussed here.
Nucleotide Competitive Inhibitors
DENV and YFV NS3hel have been co-crystallized in the
presence of a bound nucleotide (Table 1). Both structures
show a high similarity within their nucleotide binding site.
The structure of DENV NS3hel co-crystallized with the ATP
analog, ADPNP, is represented in Fig. (3) [25]. The ATP
binding site is located at the interface between subdomain 1
and subdomain 2 (Fig. 3A). A key feature of the interaction
between ADPNP and DENV NS3hel is that the adenine ring
points toward the solvent and there is little interaction with
the protein. This binding mode is also observed with ADP
co-crystallized with YFV NS3hel [43]. The main interactions
between the nucleotide/metal cofactor and the protein take
place in a narrow pocket surrounding the phosphate groups
of the ATP analog. Most of the interactions occurring in this
area are ionic/electrostatic interactions (Fig. 3B). Altogether
these structural features are not very appealing for drug dis-
covery. In an aqueous environment, hydrophobic (or apolar)
interactions are very favorable because of the changes in
solvation occurring upon the interaction between two hydro-
philic partners. It has been reported that the strength of the
interaction between a ligand and its receptor increases with
the apolar surface buried upon the interaction [44, 45]. Even
if some enhancements in potency can be made by creating
electrostatic/ionic interactions; apolar regions involved in
making the contact surface usually contribute to a large part
of the binding energy of an interaction between low molecu-
lar weight inhibitors and their receptors. For example, it has
been possible to synthesize potent low molecular weight
inhibitors of protein kinases because the adenine ring of ATP
is buried in a cavity where several hydrophobic contacts can
be created between the inhibitor and the protein. On the basis
of the available NS3hel structures, it seems that it will be
very difficult to design such inhibitors for NS3hel proteins.
There is also some intriguing experimental evidence
showing that inhibition of the ATPase activity of NS3hel
NS3 Helicases as Drug Targets Current Chemical Biology, 2009, Vol. 3, No. 3 339

might not lead to the abrogation of its helicase activity. For
example, it has been shown that the Met283Phe, His293Ala,
Thr324Ala and Cys292Gly HCV NS3hel mutant proteins
have a strongly reduced ATPase activity, while preserving a
substantial helicase activity [46-48]. Similar findings have
been observed with the DENV enzyme where the Arg460Ala
and Arg463Ala mutant proteins show reduced ATPase activ-
ity but helicase activity similar to wild-type enzyme [49].
The corresponding mutations in the JEV NS3 (Arg461Ala
and Arg464Ala) [24] and the HCV NS3 (Arg464Ala and
Arg467Lys) [46] proteins do not lead to the same results.
These mutations abolish both the ATPase and the helicase
activity of the JEV and HCV NS3 proteins. In addition, to
the results obtained with mutated proteins, it has been re-
ported that the HCV NS3hel protein has a strand-separating
activity in the absence of ATP [50]. Altogether the findings
suggest that NS3 proteins with weak (if any) ATPase activity
are still able to show good helicase activity. On the basis of
these observations, it was proposed that inhibiting the AT-
Pase activity of the NS3 enzymes may not be the right strat-
egy for drug discovery [8, 48]. However the Met283Phe mu-
tant protein, which shows a reduced ATPase activity (at
24°C) and an increased helicase activity (at 37°C) in bio-
chemical assays, has impaired functions in in vivo experi-
ments (at 37°C) [47]. Therefore the helicase activity detected
in the biochemical assays may not be sufficient in vivo, sug-
gesting that a certain level of ATPase activity is required in
vivo for full activity of the NS3 protein.
Nucleic acid Competitive Inhibitors
Different structures of NS3hel in complex with NA sub-
strates have been determined (Table 1). The bound NA is
located in a cleft formed at the interface between [subdomain
1 - subdomain 2] and subdomain 3 (Fig. 4A). Some differ-
ences exist in the interaction between the NA and HCV or
DENV NS3hel (see Luo et al. for a discussion [31]). The NA
binding site is not surprisingly larger than the ATP binding
site, and therefore low molecular weight compounds could
be easily docked into it. Looking in more detail at the inter-
actions between the NA and NS3hel reveals that the NA in-
teracts with the protein mainly via its phospho-sugar back-
bone (residues colored as white and grey sticks on Fig. 4B).
There are few contacts with the more hydrophobic parts (the
bases) of the NA and the protein (residues colored as black
lines on Fig. 4B). The only region of the DENV NS3hel with
a small hydrophobic patch formed by residues of the protein
is located near the 5’ end of the NA. However this region
does not form a defined area where inhibitors could bind
strongly. One marked difference between HCV and DENV
NS3hel is the presence of a tryptophan residue – Trp501 – at
one end of the NA binding site of the HCV protein [51]. This
aromatic amino acid shows stacking interactions with one
base of the bound nucleotide. The presence of such a residue
in the binding site could be exploited to design inhibitors that
bind to Trp501 and make additional contacts with the sur-
rounding residues. However the region around Trp501 is not
very favorable for drug discovery, and it might be difficult to
create additional interactions to enhance the potency of in-
hibitors interacting with Trp501. A low molecular weight
inhibitor of HCV NS3hel, QU663, has recently been de-
scribed [52], and docking studies suggest that it could bind
to that region of the protein. QU663 potency is low, 0.75

M, and it remains to be demonstrated if more potent ana-
logs can be designed. Furthermore the binding mode of this
molecule has to be confirmed by further structural studies.
Even if the finding of molecules such as QU663 creates
some hopes, the NA binding site of NS3hel proteins is not
very attractive for drug discovery. Potent compounds bind-
ing to this region of NS3 proteins might be very challenging,
because few hydrophobic residues are present in the cleft
where the NA binds.
It should be noted that other helicases do not interact
with NAs in the same way as NS3 enzymes. For example,
the DNA helicase PcrA makes many contacts with the bases
and shows few interactions with the phospho-sugar back-
bone [53]. Hydrophobic residues from PcrA are located
around bases, creating some kind of discrete binding pockets
where inhibitors could bind. This situation is more favorable
for drug discovery, and inhibitors binding to these pockets
have been identified [54].

Fig. (4). Nucleic acid binding site of DENV NS3hel. A. Solid ribbon representation of the DENV NS3hel protein. Subdomain 1, subdomain
2 are represented in white and subdomain 3 in black. The bound nucleic acid is schematically represented in grey. B. Interaction between the
bound NA and DENV NS3hel. The residues interacting with the phosphate groups, the sugar moieties or the bases are white sticks, grey
sticks and black lines, respectively. The bound nucleotide is represented in black sticks. For clarity neither the hydrogen bonds nor the name
of the residues are indicated. The structure used to make this representation is 2jlv [25].
340 Current Chemical Biology, 2009, Vol. 3, No. 3
Nucleotide and Nucleic Acid Non-Competitive Inhibitors
Since both the ATP and the NA binding sites show low
druggability, an alternative strategy is to identify compounds
that bind outside of these regions and inhibit NS3hel by an
“allosteric” mechanism. Since helicases are flexible proteins,
they can take different conformations during the catalytic
cycle, and one can imagine identifying compounds that
block them in one of these conformations. For example, one
might consider molecules binding to NS3hel in its apo con-
formation. Such inhibitors would then prevent the enzyme
from changing its conformation and consequently interacting
with its substrate(s). In this case, screens could be designed
so that the inhibitors are pre-incubated with the apo enzyme
and then the catalytic reaction initiated by the addition of the
substrates. Alternatively, miniaturized thermal denaturation
methods could be used to identify compounds that bind to
the apo enzyme [55]. Since NS3 and NS3hel have distinct
kinetic properties (see above) it would be more reasonable to
run these screens with the full-length protein and not only
with the isolated helicase domain.
The question is: are there druggable pockets/cavities at
the surface of the helicase domain of NS3 where compounds
could bind and inhibit the enzyme? Pockets/cavities are often
present at the surface of proteins, but not all of them are
druggable and, even if it is possible to identify a compound
that binds to one of them, it may not necessarily affect the
catalytic activity of the targeted protein. Several computer
programs enable pockets/cavities to be identified at the sur-
face of proteins. Using such tools, it is possible to find some
pockets/cavities at the surface of NS3 (see for example Fig.
5). It remains to be established if they are suitable binding
sites for low molecular compounds and if their interaction
with these pockets will abolish NS3 helicase activity.
Can this type of inhibitor be identified for RNA heli-
cases? Every protein is different, and a general rule cannot
be established. However it has been shown that this strategy
is applicable to the DEAD box RNA helicase eIF4A. Hip-
puristanol, a coral-derived natural compound, inhibits eIF4A
Fig. (5). Pockets/cavities present at the surface of the DENV NS3
protein. The helicase and the protease domains are colored in white
and grey, respectively. Pockets/cavities predicted using the Pocket
Finder software (Bioinformatics group – Leeds University) are
shown as black areas. Pocket Finder is based on the Ligsite algo-
rithm [63]. The structure used to make the figure is 2vbc [31].
Patrick Chène
upon binding to a pocket outside the ATP and NA binding
sites [12]. Strategies aimed at identifying nucleotide and/or
nucleic acid non-competitive inhibitors are complex and
require the combined forces of enzymology, biophysics and
structural biology to guide the research effort in medicinal
chemistry.
CONCLUSION
Hepaciviruses and flaviviruses are at the origin of various
human diseases and major efforts are being made to identify
medicines that will be effective in the treatment of these vi-
ruses. Identifying inhibitors of NS3 helicase activity is one
of the drug discovery strategies currently under investiga-
tion. The helicase activity of NS3 is important for viral in-
fection, making NS3hel an attractive target for drug discov-
ery. Helicases are new drug targets, and little is known about
their druggability. Using the structural information available,
three different possibilities were evaluated for identifying
inhibitors of NS3hel: the design of nucleotide competitive
inhibitors, nucleic acid competitive inhibitors and nucleotide
or nucleic acid non-competitive inhibitors. The study of the
structure of ATP and NA binding sites shows that these are
not very attractive targets for drug discovery. This suggests
that it might be very difficult to design potent low molecular
weight inhibitors targeting these regions of NS3hel proteins.
It is more difficult to draw a conclusion on the third strategy,
since some pockets/cavities are present at the surface of the
NS3 protein. Careful studies should be carried out to deter-
mine whether some of these pockets/cavities play a role in
the function of the protein. This will help in deciding
whether this strategy can be pursued. But it should be borne
in mind that, even if these pockets are important for the ac-
tivity of the protein, they should also be suitable for drug
discovery, otherwise it will not be possible to obtain potent
inhibitors. Finally, because the helicase activity is modulated
by the protease domain and other viral proteins, the design of
screening assays using the isolated kinase domain might not
be the best strategy. Assays using full-length NS3 and if pos-
sible some of its interacting partners might be a better way
for the identification of NS3hel inhibitors.
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Accepted: May 25, 2009