Cooperative Binding of the Cationic Porphyrin Tris-T4 Enhances Catalytic Activity of 20S Proteasome Unveiling a Complex Distribution of Functional States
Abstract
:1. Introduction
2. Results
2.1. Modulation by Tris-T4 of the Steady-State Chymotryptic-Like Activity of 20S Proteasome
2.2. NMR Analysis of Tris-T4/20S Proteasome Complexes
2.3. In Silico Studies
2.3.1. Ligand Analysis
2.3.2. Docking Studies on the Tris-T4-h20S Interaction
2.3.3. Analysis of the Tris-T4-Induced h20S Conformational Changes
3. Discussion
4. Materials and Methods
4.1. Chemicals
4.2. 20S Proteasomal Enzyme Assays and Analysis of Kinetic Data
4.3. NMR Measurements
4.4. Molecular Modelling
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Notes
Abbreviations
UPS | ubiquitin-proteasome system |
RPs | regulatory particles |
CP | core particle |
PCD | protein conformational disorders |
ChT-L | chymotrypsin-like |
T-L | trypsin-like |
C-L | caspase-like |
IDP | intrinsically disordered proteins |
h20S | human 20S |
y20S | yeast 20S |
SARs | structure–activity relationships |
STD | saturation transfer difference |
WL | WaterLogsy |
SA | simulated annealing |
MM | molecular mechanics |
RMSD | root mean square deviation |
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Human | Yeast | α3ΔN | |
---|---|---|---|
kcatA (s−1) | 11.5 ± 1.3 | 0.95 ± 0.15 | 49.5 ± 5.6 |
kcatB (s−1) | 0.32 ± 0.05 | 0.17 ± 0.03 | 49.5 ± 5.6 |
KmA (M) | 1.95(± 0.37) × 10−3 | 1.17(± 0.22) × 10−4 | 4.1(± 0.6) × 10−3 |
KmB (M) | 1.5(± 0.3) × 10−4 | 9.04(± 1.82) × 10−6 | 1.07(± 0.27) × 10−2 |
KAU(M−1) | 2.0(± 0.3) × 104 | 3.0(± 0.5) × 105 | 3.0(± 0.5) × 105 |
KBU(M−1) | 8.2(± 1.2) × 107 | 9.0(± 1.8) × 107 | 9.0(± 1.8) × 107 |
KAL(M−1) | 1.2(± 0.3) × 104 | 1.2(± 0.3) × 104 | 1.2(± 0.4) × 105 |
KBL (M−1) | 1.35(± 0.28) × 108 | 1.3(± 0.3) × 108 | 1.3(± 0.3) × 108 |
KBI (M−1) | - | 2.5(± 0.4) × 106 | 2.5(± 0.4) × 106 |
LU | 3.0(± 0.5) × 10−6 (3) | 6.0(± 1.0) × 10−4 (2) | 3.9(± 0.6) × 10−2 |
LL | 3.9(± 0.6) × 10−5 (3) | 7.8(± 1.5) × 10−3 (2) | 1.5(± 1.0) × 10−2 |
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Santoro, A.M.; D’Urso, A.; Cunsolo, A.; Milardi, D.; Purrello, R.; Sbardella, D.; Tundo, G.R.; Diana, D.; Fattorusso, R.; Dato, A.D.; et al. Cooperative Binding of the Cationic Porphyrin Tris-T4 Enhances Catalytic Activity of 20S Proteasome Unveiling a Complex Distribution of Functional States. Int. J. Mol. Sci. 2020, 21, 7190. https://doi.org/10.3390/ijms21197190
Santoro AM, D’Urso A, Cunsolo A, Milardi D, Purrello R, Sbardella D, Tundo GR, Diana D, Fattorusso R, Dato AD, et al. Cooperative Binding of the Cationic Porphyrin Tris-T4 Enhances Catalytic Activity of 20S Proteasome Unveiling a Complex Distribution of Functional States. International Journal of Molecular Sciences. 2020; 21(19):7190. https://doi.org/10.3390/ijms21197190
Chicago/Turabian StyleSantoro, Anna Maria, Alessandro D’Urso, Alessandra Cunsolo, Danilo Milardi, Roberto Purrello, Diego Sbardella, Grazia R. Tundo, Donatella Diana, Roberto Fattorusso, Antonio Di Dato, and et al. 2020. "Cooperative Binding of the Cationic Porphyrin Tris-T4 Enhances Catalytic Activity of 20S Proteasome Unveiling a Complex Distribution of Functional States" International Journal of Molecular Sciences 21, no. 19: 7190. https://doi.org/10.3390/ijms21197190