Clade-Specific Alterations within the HIV-1 Capsid Protein with Implications for Nuclear Translocation
Abstract
1. Introduction
2. Materials and Methods
2.1. Alignment of CA Sequences from Diverse Clades
2.2. Construction of Monomeric CA Expression Vectors from Diverse Clades
2.3. Overproduction and Purification of Monomeric CA Proteins
2.4. In Vitro Assembly Assay
2.5. Modeling and Generation of the HIV-1 CA Proteins from Diverse Clades
2.6. Overproduction and Purification of Hexameric CA Protein
2.7. Peptides
2.8. SPR Characterization
2.9. Docking Calculations for PF-74
2.10. Docking Calculations for NUP-153
2.11. Interface and Interprotomer Pocket Analysis of the HIV-1 CA Hexamer
3. Results
3.1. HIV-1 CA Protein Conservation
3.2. Overproduction and Purification of CA Monomeric Proteins from Diverse Clades
3.3. In Vitro Assembly of CA Monomeric Proteins from Diverse Clades
3.4. Overproduction and Purification of CA Hexameric Proteins from Diverse Clades
3.5. Altered Substrate Recognition across Hexameric CA Proteins from Diverse Clades
3.6. Modeling of Non-Subtype-B CA Proteins—Implications for Assembly
3.7. Modeling of Non-Subtype-B CA Proteins—Implications for PF-74 and NUP-153 Binding
4. Discussion
5. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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HIV-1 Clade | Initial Velocity Apo (AU/min) | r2 | Initial Velocity PF-74 (AU/min) | r2 |
---|---|---|---|---|
A1 (92UG037.1) | 0.022 ± 0.003 | 0.966 | 0.034 ± 0.001 | 0.996 |
A2 (CDKTB48) | 0.013 ± 0.000 | 0.990 | 0.017 ± 0.002 | 0.983 |
B (NL4-3) | 0.016 ± 0.003 | 0.945 | 0.043 ± 0.009 | 0.925 |
C (92BR025.8) | 0.011 ± 0.000 | 0.929 | 0.039 ± 0.007 | 0.930 |
D (94UG114.1) | 0.015 ± 0.002 | 0.954 | 0.050 ± 0.008 | 0.955 |
HIV-1 Clade | KD [nM] Equilibrium | ±SD | kon [M−1s−1] | ±SD | koff [s−1] | ±SD | KD [nM] Kinetic/Global | ±SD |
---|---|---|---|---|---|---|---|---|
A1 | 716.3 | 66.1 | 6.15 × 105 | 1.80 × 105 | 6.47 × 10−1 | 2.82 × 10−1 | 946.5 | 313.5 |
A2 | 138.0 | 16.5 | 6.69 × 105 | 1.78 × 105 | 9.16 × 10−2 | 1.42 × 10−2 | 139.3 | 14.6 |
B (NL4.3) | 47.0 | 0.6 | 7.56 × 105 | 4.55 × 104 | 3.63 × 10−2 | 6.92 × 10−3 | 47.9 | 1.2 |
C | 353.7 | 10.0 | 2.28 × 105 | 2.20 × 104 | 7.54 × 10−2 | 6.92 × 10−3 | 330.0 | 14.7 |
D | 51.0 | 1.3 | 7.73 × 105 | 4.01 × 104 | 3.95 × 10−2 | 5.25 × 10−3 | 51.0 | 4.1 |
HIV-1 Clade | KD [μM] Equilibrium | ±SD | kon [M−1s−1] | ±SD | koff [s−1] | ±SD | KD [μM] Kinetic/Global | ±SD |
---|---|---|---|---|---|---|---|---|
A1 | 181.0 | 55.9 | 1.85 × 103 | 9.29 × 101 | 4.77 × 10−1 | 3.27 × 10−2 | 257.3 | 7.8 |
A2 | 197.0 | 35.4 | 3.68 × 103 | 1.33 × 103 | 7.46 × 10−1 | 1.73 × 10−1 | 209.7 | 28.2 |
B (NL4.3) | 93.8 | 3.3 | 5.96 × 103 | 1.28 × 103 | 5.89 × 10−1 | 1.28 × 10−1 | 98.6 | 4.7 |
C | 104.3 | 3.8 | 5.29 × 103 | 1.21 × 103 | 5.70 × 10−1 | 1.28 × 10−1 | 107.7 | 1.5 |
D | 70.0 | 0.7 | 7.06 × 103 | 1.14 × 103 | 5.13 × 10−1 | 8.05 × 10−2 | 72.8 | 0.9 |
HIV-1 Clade | KD [μM] Equilibrium | ±SD | kon [M−1s−1] | ±SD | koff [s−1] | ±SD | KD [μM] Kinetic/Global | ±SD |
---|---|---|---|---|---|---|---|---|
A1 | N.B. | N.B. | N.B. | N.B. | N.B. | N.B. | N.B. | N.B. |
A2 | 443.3 | 38.1 | 130 × 103 | 3.75 × 102 | 9.60 × 10−1 | 2.13 × 10−1 | 746.3 | 48.5 |
B (NL4.3) | 380.7 | 34.6 | 1.57 × 103 | 3.60 × 102 | 7.63 × 10−1 | 7.35 × 10−2 | 499.3 | 94.1 |
C | 503.0 | 46.9 | 9.33 × 102 | 4.31 × 102 | 7.47 × 10−1 | 7.35 × 10−2 | 881.0 | 267.2 |
D | 308.7 | 42.8 | 1.89 × 103 | 2.91 × 102 | 6.86 × 10−1 | 1.35 × 10−1 | 367.7 | 83.0 |
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Dick, A.; Meuser, M.E.; Cocklin, S. Clade-Specific Alterations within the HIV-1 Capsid Protein with Implications for Nuclear Translocation. Biomolecules 2022, 12, 695. https://doi.org/10.3390/biom12050695
Dick A, Meuser ME, Cocklin S. Clade-Specific Alterations within the HIV-1 Capsid Protein with Implications for Nuclear Translocation. Biomolecules. 2022; 12(5):695. https://doi.org/10.3390/biom12050695
Chicago/Turabian StyleDick, Alexej, Megan E. Meuser, and Simon Cocklin. 2022. "Clade-Specific Alterations within the HIV-1 Capsid Protein with Implications for Nuclear Translocation" Biomolecules 12, no. 5: 695. https://doi.org/10.3390/biom12050695
APA StyleDick, A., Meuser, M. E., & Cocklin, S. (2022). Clade-Specific Alterations within the HIV-1 Capsid Protein with Implications for Nuclear Translocation. Biomolecules, 12(5), 695. https://doi.org/10.3390/biom12050695