Transcript (π(I 2 )→π (nucleot.))

Scientific Centre of Anti-Infectious Drugs
Kazakhstan
A Quantum-Chemical Model of the
Inhibition HIV-1 intergrase action by
Iodine Complex Compounds
Gulnara Yuldasheva, Aleksandr Ilin
Preintegration complex
HIV integrase inhibitors have a high therapeutic effect for two reasons.
First, integrase is one of the key participants in the virus replication cycle.
Second, integrase has no cellular equivalent and, hence, the suppression of its
activity should not disturb normal cellular metabolism processes.
Sen, S., Mathur, A.G., Gupta, R.M., Kapila, K., and Chopra, G.S. (2008)
Recent Pat Antiinfect Drug Recent Pat Antiinfect Drug Discov., 3(3), 199-205.
In the paper the authors offer a comparison of the inhibiting activity of a
series of antiretroviral agents towards HIV-1 integrase and the in vitroisolated PIC. The results of the study show that inhibitors active towards the
integrase enzyme may not be active towards the PIC. The capability to
inactivate a PIC was exhibited only by three antiretroviral agents:
quinalizarin, purpurin, and alizarin.
Farnet, C.M., Wang, B., Lipford, J.R., and Bushman, F.D. (1996)
Differential Inhibition of HIV -1 Preintegration Complexes and Purified
Integrase Protein by Small Molecules. Proc.Natl. Acad. Sci. USA, 93, 97429747.
We propose a model of inhibition of HIV-1 intergrase action where the
active center of those drugs inhibits the integrase catalytic center inside the
PIC.
Armenicum
Well-known are quite a number of iodine-polymer complexes that possess a broad
spectrum of antimicrobic and antiviral action including human immunodeficiency virus
(HIV).
Among them is Armenicum which is a drug to treat HIV infection.
Patent Pub.No. WO/2001/078751 International Application No.: PCT/AM2000/000002,
International Filling Date 24.11.2000.
Recently a new anti-infective drug with anti-HIV action has been patented. At present
the new drug (FS-1) is in the third clinical trial and for that reason has no name yet.
Patent KZ (A) №15116, 15.12.2004, bull 12.
Armenicum is composed of the LiI5--dextrin complex. The FS-1 contains polypeptides
along with the LiI5 --dextrin complex.
Gulnara A. Yuldasheva, Georgii M. Zhidomirov, and Aleksandr I. Ilin «Effect of Organic
Ligands with Conjugated π-Bonds upon the Structure of Iodine--Dextrin Complexes.»
Biotechnology and Apply Biochemistry 2012 Jan-Feb, 59(1), P.29-34.
We managed to get the this electronic structure of I2 only by using the quantumchemical calculations. However, using UV-IR – spectroscopy and the quantum-chemical
method DFT-B3PW91/midi we researched the water-glycine – KI3 - LiCl – ethanol
system as a model of the FS-1 active centre.
[Gulnara A. Yuldasheva, Georgii M. Zhidomirov, Jerzy Leszczynski, Aleksandr I. Ilin «The
effect of the amphoteric properties of amino acids in the zwitterionic form on the structure
of iodine complex compounds in aqueous solutions containing halogenides of alkaline
metals and amino acids» Journal of Molecular Structure 1033 (2013) 321–330.
Active center of FS-1
∆E=-16,60
∆E= -6,25
∆E=-14,26
∆E=-24,25
Energy of formation (∆E, kkal/mol) active complex (amine acid residues, ethanol, molecular
iodine LiCl). Red balls - oxygen atoms, blue - carbon, nitrogen – dark blue, violet-iodine,
lithium-brown, chlorine - yellow.
Model of interaction of the active complex Armenicum and
FC-1 with nucleotides of HIV DNA
red balls - oxygen atoms, blue - carbon, nitrogen-dark blue, violet-iodine, yellow –
chlorine, lithium-white.
UV – spectrum of system (a-c)
1,1
199,43nm 0,94676A
1,0
0,9
0,8
256,56nm; 0,61A
0,7
A
0,6
0,5
0,4
0,3
0,2
0,1
0,0
200
300
Description
Name
AGA 2.Sample Н2О
400
nm
500
600
700
System (a)
AGA triplet
1,2
1,1
1,0
223,08nm; 0,59A
0,9
0,8
A
0,7
256,43nm 0,14933A
0,6
0,5
System (b) FS-1
0,4
460,99nm; 0,01A
355,15nm; 0,01A
0,3
0,2
0,1
0,0
200
300
Name
R10.Sample
Description
400
500
600
nm
Н2О
System (c) AGA triplet+FS-1
700
Table 1. Theoretical wavelengths (λ, nm) of electronic transitions in iodine
complex with adenosine and lithium halogenides (TD-DFT/B3PW91/midi)
λ theor.
206,51
1A→1A (π(I
1A→1A (π
2)→π
(nucleot.))
(nucleot.)→π (nucleot.))
210,20
1A→1A (π(I
218,63
1A→1A (π
2)→π (nucleot.))
1A→1A ((Cl—Li--I)→ π(I ))
2
(nucleot.)→ π(I2))
π(I2))
1A→1A ((N—I-I)→
(nucleot.)→ π(I2))
226,57
1A→1A (π
234,55
1A→1A (π(I
244,34
2)→ π(I2))
1A→1A (π
(nucleot.)→ π(I2))
1A→1A (π
(nucleot.)→π (nucleot.))
Table 2. Theoretical wavelengths (λ, nm) of electronic transitions in iodine
complex with guanosine and lithium halogenides (TD-DFT/B3PW91/midi)
λ theor.
215,09
1A→1A (π(I
2)→
π(I2))
233,71
1A→1A (π(I
2)→π
244,34
1A→1A (π(I
2)→
(nucleot.))
π(I2))
Mechanism of Inhibition of the catalytic center of the
HIV-1 Integrase
The structure of the catalytic domain of the HIV-1 integrase is determined
by the X-ray structural analysis. According to the data obtained, the catalytic
domain of the enzyme in the crystal forms a spherical dimer with each
monomer forming a semi-sphere. The three amino acid residues - Asp64,
Asp116 and Glu152 closely located in the tertiary structure of the catalytic
domain – form its active catalytic center with the active centers of each
integrase monomer located on the opposite sides of the dimer sphere 35Å
apart.
The X-ray structural analysis data clearly show one Mg2+ ion coordinated
by Asp64 и Asp116 and two water molecules.
Dyda, F., Hickman, A.B., Jenkins, T.M., Engelman, A., Craigie, R., and Davies D.R.
(1994) Crystal Structure of Catalytic Domain of HIV-1 Integrase Similarity to Other
Polynucleotidyl Transferases. Science, 266, 1981-1986.
Goldgur, Y., Dyda, F., Hickman, A.B., Jenkins, T.M., Craigie, R., and Davies, D.R.
(1998) Three New Structures of the Core Domain of HIV-1 Integrase: An Active Site that
Binds Magnesium. Proc.Natl.Acid.Sci USA, 95(16), 9150-9154.
Based
on
data
regarding
the
action
mechanism
of
nucleotidyltransferases, referred to the same family as the HIV-1
integrase, it was suggested in article that two ions may take part in the
act of catalysis, but, due to a higher conformational mobility of the
catalytic enzyme, they are not coordinately bound prior to being bound
with the virus DNA.
With the use of the molecular dynamics method it was shown that the
interaction with the virus DNA involves the integration of two Mg2+ ions
into a stable bi-nuclear structure where the amino acid residue Glu152
simultaneously coordinates the two Mg2+ ions, while Asp64 and Asp116
interact only with one of the Mg2+ ions.
Chen, X., Tsiang, M., Yu, F., Hung, M., Jones, G.S., Zeynalzadegan, A., Qi, X.,
Jin, H., Kim, C.U., Swaminathan, S., and Chen J.M. (2008) Modeling, Analysis, and
Validation of a Novel HIV Integrase Structure Provide Insights into the Binding
Modes of Potent Integrase Inhibitors. J.Mol.Biol., 380, 504-519.
Nucleoprotein complex formed by nucleotide of virus DNA,
active center of FS-1, active catalytic center.
I, ∆E = -22,3
III, ∆E = -28,3
II, ∆E = -19,7
IV, ∆E = -20,3
Energy of formation (∆E, kkal/mol) of nucleoprotein complex. Red balls - oxygen atoms,
blue - carbon, nitrogen – dark blue, violet-iodine, lithium-brown, chlorine – yellow,
magnesium-green.
Experimental Study of FS-1 Substance in Relation to HIV-1 Virus
A study of the antiviral effect of FS-1 substance in relation to HIV-1 virus was
conducted. The study was conducted using a line of human lymphoblastoid cells (MT2). Line H9 / HTLV-IIIB of human cells chronically infected with HIV-1LAI was used
a source of HIV-1. Anti-HIV effect of FS-1 was determined based on the level of protein
p24 and reverse transcriptase activity.
Table 3 shows the results of a study of viral activity of the HIV-1 stock solution in
MT-2 cell culture.
Table 3. Results of Infectivity in terms of Cytopathic Effect, Level of p24 Antigen
and RT-Activity of HIV-1 in MT-2 Cells.
Dilution of
Stock Solution
MTT
A450
P24
A450
RT
A405
Volume of
Liquid
% Living Cells
100
0,506
2,907
1,300
0,05ml
0,506/1,157=44
10-1
0,918
2,699
1,724
0,05ml
0,918/1,157=79
10-2
0,908
2,771
1,085
0,05ml
0,908/1,157=78
10-3
1,012
0,746
1,010
0,05ml
1.012/1,157=87
10-4
1.000
0,423
0,954
0,05ml
1,000/1.157=86
Experiment Design:
200 l (80,000-100,000) of uninfected cells МТ-2 were placed into 96-well plates.
24 hours later they were infected with HIV-1 (Stock 13) and were put into an oven
for 1 hour at 37 °С. Then the supernatant was removed and the test substance at
various concentrations was introduced at the rate of 6 repetitions per each
concentration. A number of uninfected cells were used as negative controls, while a
number of virus-infected cells not treated with the test substance were used as
positive controls. Percentage of survived cells was calculated after the computation
of the average value of optical density of the experiment.
The results of the experiment have shown that FS-1 has a clear cut anti-HIV
activity in MT-2 cell culture.
The experiment was conducted at The National Centre for Infectious Diseases
of the Republic of Bulgaria under the guidance of Professor R. Argirova
Table 4 Anti-HIV Action of FS-1 as shown by the Level of Reverse Transcriptase
Activity
Experimen
t Date
FS-Dilution
1:4,00
% of Killed
Cells
% inhibition of
HIV Infection in
Terms of RT
Activity
1:8,000
(MNC)
1:16,000
1:8,000
(1/2MNC)
(1/4MNC)
18.12.07
3
35
50
-
21.0108
7
39
55
90
25.01.08
10
45
51
83
18.12.07
-
-
-
-
21.0108
81
60
50
8
25.01.08
92
65
54
At 1:4,000 dilution the activity of reverse transcriptase was 92% inhibited after 7 days
Conclusions
The computation results show that drug FS-1 may be
referred to compounds inhibiting the catalytic center of
the HIV-1 integrase.
Our research have shown that active center of FS-1
prevents the formation of PIC and inhibits the HIV-1
integrase inside the nucleoprotein complex where the
binuclear I2LiOCOMg complex interacts both with the
virus DNA and the active center of the catalytic domain
of the HIV-1 integrase.
The results of the experiment have shown that FS-1
has a clear cut anti-HIV activity in MT-2 cell culture.