Supplementary data
The complex and specific pMHC interactions with diverse HIV-1
TCR clonotypes reveal a structural basis for alterations in CTL
function
Zhen Xia1,ǂ, Huabiao Chen2,ǂ, Seung-gu Kang1, Tien Huynh1, Justin W. Fang2, Pedro A.
Lamothe2, Bruce D. Walker2,3*, and Ruhong Zhou1,4*
1Computational
Biology Center, IBM Thomas J. Watson Research Center, Yorktown
Heights, New York, USA
2Ragon
Institute of Massachusetts General Hospital, Massachusetts Institute of
Technology, and Harvard University, Cambridge, Massachusetts, USA
3Howard
Hughes Medical Institute, Chevy Chase, Maryland, USA
4Department
ǂThese
of Chemistry, Columbia University, New York, New York, USA
authors contributed equally to this work.
*Correspondence should be addressed to B.D.W. (bwalker@partners.org) or R.Z.
(ruhongz@us.ibm.com)
Note 1 for the clonetype B6: Our FEP calculations also support this relatively weak
antiviral efficacy of the clonotype B6, with binding affinities severely decreased by
either the R2T (ΔΔG = 6.89 ± 1.20 kcal/mol) or R2TL6M (ΔΔG = 4.84 ± 0.86
kcal/mol) mutantations, largely due to destruction of the hydrogen-bond network at
the N-terminus of KK10 peptide (Supplementary Fig. 4). The more conservative
mutation L6M slightly increases the binding affinity by ΔΔG = -1.37 ± 0.82 kcal/mol
(Supplementary Table 1), in which an additional hydrogen bond is formed
between the KK10 peptide (Asn9) and the CDR3β of the TCR (Gly100) in the L6M
mutant (Supplementary Fig. 4a). However, the overall binding may not be
sufficiently enhanced even with this favorable mutation due to the absense of a
compact hydrophobic core inherited from the WT KK10 peptide, likely resulting in
the weak antiviral efficacy overall.
Note 2 for the clonetype B6: Similarly, the binding affinity changes of alanine
mutants in B6 clone are very close to those in vivo mutants (Supplementary Fig. 7).
Again, L6A shows a slightly stronger binding (ΔΔG = -1.62 ± 0.67 kcal/mol), which is
partly due to the same new hydrogen bond formed between the KK10 peptide
(Asn9) and the CDR3β of the TCR (Gly100) as in L6M mutant, and partly due to an
“induced-fit” for neighboring hydrophobic residues Ile5 in KK10 peptide and Val97
at the CDR3α of the TCR because of the relatively smaller size of alanine
(Supplementary Fig. 7a).
Note 3 for free energy perturbation protocol details
The binding affinity changes, due to antigenic variations, between the TCR and HLAKK10 complex were estimated by the free energy perturbation (FEP) method (111). The binding free energy change G due to a mutation in the KK10 of the HLAKK10 binary complex can then be calculated as,
G  kT ln exp(  [V (   )  V ( )]
G   G

(1)

(2)
where V(λ) = (1- λ) V1 + λ V2, and V1 represents the potential energy of the wild-type,
and V2 represents the potential energy of the mutant. The FEP parameter λ changes
from 0 (V1) to 1 (V2) when the system changes from the wild-type to the mutant, and
<…> λ represents the ensemble average at potential V(λ).
In general, it is difficult to directly calculate the absolute binding affinity change GA
for the binding process between two interacting surfaces, like in the interface of the
TCR and HLA-KK10 binary complex, due to the long time scale and complicated
binding process. However, we can avoid this problem by designing a
thermodynamic cycle to calculate a relative binding free energy change, i.e., GAB.
Instead of calculating the difficult direct binding energies GA and GB, we calculate
the free energy changes for the same mutation in both the bound state (HLA-KK10TCR 3-way binding complex, G1) and the free state (HLA-KK10 binary complex,
G2). Within a complete thermodynamic cycle, the total free energy change should
be zero, which gives the relative binding affinity due to the mutation from A→ B as:
G
bind
 G  G  G  G
B
A
1
2
(3)
For each mutation, at least five independent runs starting from different initial
configurations (taken from the molecular dynamic simulations) are performed for
better sampling. The simulation time for each FEP simulation is 6.0 ns. Larger
window sizes and longer simulation durations have also been tested in our previous
studies, and we found that the current protocol gives us a reasonable convergence
in the final binding affinities (9, 10). Therefore, at least 60 ns (6.0-ns  5-runs  2states) simulation time was generated for each mutation, which is much longer than
most FEP calculations currently reported in the literature.
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Tables
Supplementary Table 1:
Align TCR α chain segment and β chain segment to known structures
Supplementary Table 2:
Sequence identities among clonotypes B3, B5, and B6
Clones
α- chain
β- chain
B3 and B5
27%
32%
B3 and B6
90%
24%
B5 and B6
29%
31%
Supplementary Table 3: The FEP simulation results for the TCR-KK10 binding free
energy change due to the L6M, R2T, and R2TL6M mutants in viral peptide KK10a
Calculated ΔΔG (kcal/mol)
Mutants
B3 clone
B5 clone
B6 clone
L6M
0.81 ± 0.24
0.07 ± 0.30
-1.37 ± 0.82
R2T
7.88 ± 1.80
0.78 ± 1.21
6.89 ± 1.20
R2TL6M
13.17 ± 0.83
0.92 ± 0.95
4.84 ± 0.86
aA
total of five independent runs has been performed for both the bound and free states for the
standard error calculations.
Supplementary Table 4: FEP simulation results for the predicted TCR-KK10
binding free energy change due to the L6A and R2A single mutations, and R2AL6A
double mutation in viral peptide KK10a
Calculated ΔΔG (kcal/mol)
Mutants
B3 clone
B5 clone
B6 clone
L6A
0.50 ± 0.51
0.54 ± 0.61
-1.62 ± 0.67
R2A
8.13 ± 3.55
1.51 ± 2.32
7.47 ± 0.99
R2AL6A
9.71 ± 0.95
3.47 ± 0.38
5.86 ± 0.93
aA
total of five independent runs has been performed for both the bound and free states for the
standard error calculations with each running 6 ns.
Supplementary Table 5: FEP simulation results for the predicted TCR-KK10
binding free energy change due to the L6M and R2T single mutations in viral
peptide KK10. The initial wild-type structures were built based on the newly
released HLA-KK10-TCR complex structure (PDB entry: 4G8G)a
Calculated ΔΔG (kcal/mol)
Mutants
B3 clone
B5 clone
B6 clone
L6M
0.53 ± 0.25
-0.11 ± 0.33
-1.19 ± 0.29
R2T
7.93 ± 0.82
1.14 ± 0.22
7.88 ± 0.87
aA
total of five independent runs has been performed for both the bound and free states for the
standard error calculations with each running 6 ns.
Supplementary Figure legends:
Supplementary Figure 1. Comparison of the structural stability during the
simulations for WT viral peptide-HLA-TCR complexes. The plot shows The RMSD
values of the HLA B*2705-KK10-TCR 3-way binding complex. The RMSD values are
calculated by comparing each snapshot to the backbone of the starting built
structures during molecular dynamics simulations. The results are obtained from
NPT ensemble simulations (T=310 K, P=1 atm) with the simulation time of 50 ns.
Four independent repeats are performed for each clonotype. (a) Complex with B3
clone. (b) Complex with B5 clone. (c) Complex with B6 clone.
Supplementary Figure 2. Structural comparison of HLA B*2705-KK10-TCR
complexes due to in vivo occurring mutants in KK10 peptide for FW56 clone B5
(green, HLA; cyan, the HIV KK10 peptide; magenta, TCR). (a) L6M mutant. (b) R2T
mutant. (c) and (d) R2TL6M double mutant. The overall complex is represented as
cartoon and the residues at the binding site are rendered with spheres (non-polar
interactions) or sticks (polar interactions).
Supplementary Figure 3. Structural comparison of HLA B*2705-KK10-TCR
complexes due to in vivo occurring mutants in KK10 peptide for FW56 clone B6
(green, HLA; cyan, the HIV KK10 peptide; magenta, TCR). (a) L6M mutant. (b) R2T
mutant. (c) and (d) R2TL6M double mutant. The overall complex is represented as
cartoon and the residues at the binding site are rendered with spheres (non-polar
interactions) or sticks (polar interactions).
Supplementary Figure 4. Structural comparison of HLA B*2705-KK10-TCR
complexes responding to KK10 peptide variants R2A, L6A, and R2AL6A for FW56
clone B3 (green, HLA; cyan, the HIV KK10 peptide; magenta, TCR). The overall
complex is represented as cartoon and the residues at the binding site are rendered
with spheres (non-polar interactions) or sticks (polar interactions).
Supplementary Figure 5. Structural comparison of HLA B*2705-KK10-TCR
complexes responding to KK10 peptide variants R2A, L6A, and R2AL6A for FW56
clone B5 (green, HLA; cyan, the HIV KK10 peptide; magenta, TCR). The overall
complex is represented as cartoon and the residues at the binding site are rendered
with spheres (non-polar interactions) or sticks (polar interactions).
Supplementary Figure 6. Structural comparison of HLA B*2705-KK10-TCR
complexes responding to KK10 peptide variants R2A, L6A, and R2AL6A for FW56
clone B6 (green, HLA; cyan, the HIV KK10 peptide; magenta, TCR). The overall
complex is represented as cartoon and the residues at the binding site are rendered
with spheres (non-polar interactions) or sticks (polar interactions).
Supplementary Figure 7. Structural comparison of different clonotypes bound to
the KK10 peptide. The structures were built based on a new co-crystal structure
with the same HLA-KK10 but a different TCR clonotype (PDB entry: 4G8G). The
binding site and interactions at the N-terminus (a), the C-terminus (b), and the
middle region (c) of KK10 peptide are rendered with spheres (non-polar
interactions) or sticks (polar interactions) (green, HLA; cyan, HIV KK10 peptide;
magenta, TCR). The overall complexes are represented as cartoons.
Supplementary Figure 8. The salt-bridge distance between Lys1 (KK10) and Asp29
(TCR) in B3 clonotype during FEP simulations of R2T mutant. The distance was
calculated between the nitrogen atom (NZ) at Lys side-chain and the carbon atom
(CG) at Asp side-chain. The X-axis shows the FEP simulation process which
gradually perturbed Arg (λ=0) to Thr (λ =1).
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