A rational approach to a HIV-1 vaccine design - CBS

A rational approach to a HIV-1 vaccine design
Course 27685 Immunological Bioinformatics
Gustav Ahlberg, Joakim Ro-Poulsen and Snezana Djurisic
Center For Biological Sequence Analysis CBS , Technical University of Denmark DTU
Aim: To do a rational selection of peptides with a high potential as vaccine
candidates against HIV-1 in Guinea-Bissau focusing on HLA-E.
Methodology: The selection of peptides is tailored towards the prevalent HLADRB1 and HLA-E molecules present in the population in Guinea-Bissau using
bioinformatic tools. The process involved the following:
Step 1: Selecting MHC Class I/II alleles from the population.
Step 2: Predicting and selecting the peptides in the HIV 1 gag-protein
binding to the chosen alleles.
Step 3: Evaluating peptides by visualizing and reviewing mutation frequency.
Background: The human immunodeficiency virus (HIV) has been shown to abrogate
and/or down-regulate most MHC class I molecules as an evasion strategy that targets the
antiviral activity of the immune system[9,10]. Interestingly, the non-classical HLA molecule
HLA-E seems not to be affected by the viral infection[10].
HLA-E is a conserved class Ib molecule characterized by a limited polymorphism (2 coding
variants are identified to date). Its primary purpose is to bind small leader peptides of most
HLA class I molecules after which the HLA-E molecule is stabilized and migrates to the cell
surface where it interacts with receptors from natural killer (NK) cells. Recent evidence
reveals that several peptides other than MHC class I molecules encode peptides that can
bind to HLA-E including peptides derived from the HIV gag protein[8,10]. In addition, HLAE restricted CD8+ T cells have been reported against various bacterial and viral
infections[11,12].
HLA-E restricted T cell responses may be of interest for vaccine development since HLA-E
displays very limited polymorphism, and is not down-regulated by HIV-infection.
Persistent HLA-E restricted CD8+ T cell responses require assistance from CD4 + T cells
and HLA class II epitopes should also be evaluated in the context of HIV-1 vaccine design.
Step 1b: HLA Sequence Logos.
HLADRB1*1001
HLADRB1*1304
HLADRB1*0901
HLA-E*0101 and
E*0103
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Figure 1b. Three selected logos predicted via MHC Motif Viewer (upper figures)
showing the bias for peptides binding to the HLA-DRB1 molecule. Position 1, 4, 6
and 9 (the abscissa) have a high degree of information content[1,4], the most
pronounced degree found at position 1 for all three alleles. These are anchor
positions that to a high extend determines the binding of a peptide. In the depicted
logo plots, the amino acids are colored according to their properties: Acidic = red
(DE); Basic = blue (HKR); Hydrophobic = black (ACFILMPVW); Neutral = green
(GNQSTY). No information could be obtained on HLA-E with either the MHC Motif
Viewer or Syfpeithi MHC database.
Step 2a: Predicting the binding peptides.
Step 1a: Identification of the prevalent HLA-DRB1 alleles in a
Guinea-Bissau population.
Figure 2a. NetMHCpanI In-House server, predicting epitopes for HLA-
Figure 1a. The HLA-DRB1 locus has one of the highest sequence variation within the human
genome. Allelic frequencies in a Guinea Bissau population were identified using the Allele
Frequency Database[3]. Prevalent alleles were chosen to complement eachother at anchor
positions (se figure 1b), so that the alleles did not have the same binding preferences. The
alleles chosen were HLA-DRB1*09:01, HLA-DRB1*13:04, HLA-DRB1*10:01 (shown in colour).
Among these the unusual HLA-DRB1*1304 allele showed to be highly abundant covering
19.6% of the analyzed group. All together the phenotypic population coverage was calculated
to be 61%. Allel frequences for HLA-E could not be identified using the Allele frequency
Database.
Step 3a: Visualizing the protein with epitopes and amino properties.
(a) Cartoon
(b) Surface
E0101[6]. NetMHCpan identified two potential epitopes in the gag protein as
binders to HLA-E*01:01, one as a weak binder (WB) and one as strong binder
(SB).
The two epitopes (with sequence positions) were:
Weak binder:
163 FSPEVIPMF
Strong binder:
274 RMYSPTSIL
Step 2b: Epitope selection of peptides tailored to bind to
HLA-E*0101 and HLA-DRB1*1001, DRB1*0901, DRB1*1304.
Figure 2b. Seven 15mer
peptides selected for further
analysis. Four of these were
also predicted as binders for
the nanomers found with
MHCpanI (163, 272 and 273)
[7].
X represents binder and 0
represents a non binder.
Step 3b: Selected peptides are cross-reactive towards other
subtypes.
(c) Hydrophobicity
(d) Electrostatic potential
Aligning multiple gag protein sequences with blastp (AAD39400, BAF32552,
CAJ01233, ADN38206 amongst 50 others) revealed a few insimilarities in the
binding peptides[5]. These differences were shown to have little to no difference in
the predictions of netMHCpan and netMHCIIpan. Studying the prediction revealed
that most substitions
(but one, which were T-V) were of similar amino acid types and only one insertion
had been made throughout the sequence.
Results
Figure 3a. Structure of the HIV-1 gag protein and the predicted epitopes 268
GLNKIVRMYSPTSIL (blue), 131 YPIVQNIQGQMVHQA (purple), 164 SPEVIPMFSALSEGA
(orange) and 56 CRQILGQLQPSLQTG (pink). PyMol models are based on the CPHmodel
predicted from AAC82593[2,5]. The CPHmodel were predicted with 95.7% identity.
There were no clear link between the epitopes, hydrophobicity and electrostatic potential.
The resulting analysis revealed two potential 9mer epitopes found in the HIV
protein known as Gag. A BLASTp of 40 HIV 1 gag protein sequences, that were
arbitrary picked from NCBI database, also indicated that the binding properties of
the epitopes were conserved.
We found the two HLA-E binding epitopes to be located closely in the protein
structure, while the class II were more spread. There were no clear correlation
between hydrophobity or electrostatic potential and the epitopes.
The gag protein can be considered conserved compared to other proteins in the
HIV virus, a virus that displays a high genetic variability. The protein is believed
to play a crucial role in the HIV assembly process and is therefore an attractive
target in the development of antiviral drugs.
References:
[1]http:// www.syfpeithi.de/Scripts/MHCServer.dll/FindYourMotif.htm [8] Nattermann J. et al, HIV-1 infection leads to increased HLA-E expression resulting in impaired function of natural killer cells, Antivir Ther, 2005
[2]http://www.cbs.dtu.dk/services/CPHmodels-2.0/
[9] Schaefer, M.R., et al., HIV-1 Nef targets MHC-I and CD4 for degradation via a final common beta-COP-dependent pathway in T cells. PLoS Pathog, 2008. 4(8): p. e1000131
[3]http://www.allelefrequencies.net/
[10] Pietra, G., et al., The emerging role of HLA-E-restricted CD8+ T lymphocytes in the adaptive immune response to pathogens and tumors. J Biomed Biotechnol. 2010: p. 907092.
[4]http://www.cbs.dtu.dk/biotools/MHCMotifViewer/MHC_Fight.html [11] Salerno-Goncalves, R., et al., Identification of a human HLA-E-restricted CD8+ T cell subset in volunteers immunized with Salmonella enterica serovar Typhi strain Ty21a typhoid vaccine. J Immunol, 2004. 173(9): p. 5852-62.
[5]http://www.ncbi.nlm.nih.gov/
[12] Romagnani, C., et al., HLA-E-restricted recognition of human cytomegalovirus by a subset of cytolytic T lymphocytes. Hum Immunol, 2004. 65(5): p. 437-45.
[6]http://www.cbs.dtu.dk/services/NetMHCpan/
[7]http://www.cbs.dtu.dk/services/NetMHCIIpan/