Molecular Simulation

We use molecular dynamics simulations to understand how proteins move and how this plays a role in their function.  Specific examples are shown below.

 

1.  Flexibility at the immune synapse

2.  Dynamic Requirements for optimal enzyme activity

3.  Conformational dynamics of disease-causing variants of the serpin alpha-1 antitrypsin

4. Determinants of the substrate specificity of the complement initiating proteases

Epitope Flexibility and Dynamic Footprint Revealed by Molecular Dynamics of a pMHC-TCR Complex

Reboul CF, Meyer GR, Porebski BT, Borg NA, Buckle AM (2012) Epitope Flexibility and Dynamic Footprint Revealed by Molecular Dynamics of a pMHC-TCR Complex. PLoS Comput Biol 8(3): e1002404. doi:10.1371/journal.pcbi.1002404

When pathogens replicate within a host cell, their proteins are degraded into peptides, which are captured by the major histocompatibility complex (MHC) and brought to the cell surface. The peptide-MHC (pMHC) is surveyed by T cell receptors (TCRs) expressed on the surface of T cells. If the peptide is foreign, the peptide-MHC-TCR interaction initiates an immune response to eliminate the pathogen. However, the combinations of pMHC and TCRs are diverse. We ask how TCRs discriminate between structurally similar pMHCs? We address this by focusing on two MHC molecules that differ by a single change, both bind the same peptide but only one instigates a dominant immune response. Intriguingly, the single difference between the two MHCs does not alter the peptide shape nor does it contact the peptide or TCR. We examined the flexibility of the pMHC-TCR interface using molecular dynamics simu- lations. We observed differences in the peptide and TCR flexibilities that could explain their contrasting physiolo- gies, as well as clues to how the TCR moves atop the MHC in order to ‘scan’ it. Our analysis provides insight into a particular pMHC-TCR interaction not accessible using crystallographic methods, and indicate dynamics may play an influential and perhaps under-appreciated role in other pMHC-TCR systems.

 

 

Movie 1: Conformational flexibility of peptide in HLA-B*3501-LPEP and HLA-B*3508-LPEP during MD simulations. The polymorphic (R/L156) residue is shown in green. Static snapshots of the respective simulations are shown in the insets.

 


 

Movie 2: Orthogonal views of MD simulation of HLA-B*3508-LPEP SB27 TCR complex B. The MHC, LPEP epitope and TCR are shown in grey, green and yellow/cyan respectively.

 

 

 

Movie 3: Detailed view of contacts made at the interface during the MD simulation of HLA-B*3508-LPEP SB27 TCR complex B. On the left, cartoon showing hydrogen bonds and salt bridges (black dotted lines) at the interface between MHC (grey), peptide (green) and TCR (cyan and yellow); On the right, MHC surface showing the dynamic TCR footprint. The MHC residues being contacted by the TCR are coloured red. The inset shows the corresponding footprint calculated from the crystal structure.

 

 


Structural and Dynamic Requirements for Optimal Activity of the Essential Bacterial Enzyme Dihydrodipicolinate Synthase

Reboul CF, Porebski BT, Griffin MDW, Dobson RCJ, Perugini MA, Gerrard JA, and Buckle AM (2012) Structural and Dynamic Requirements for Optimal Activity of the Essential Bacterial Enzyme Dihydrodipicolinate Synthase. PLoS Comput Biol 8(6): e1002537. doi:10.1371/journal.pcbi.1002537

Enzyme function requires the specific placement of residues in the active site so that the correct chemistry is available for efficient catalysis. However, the inherent flexibility of proteins can present challenges in fulfilling these stringent requirements. We have investigated the role of flexibility in the enzyme Dihydrodipicolinate synthase (DHDPS), which in E. coli is a homotetramer consisting of a ‘dimer of dimers’, with the catalytic residues found at the tight-dimer interface. It is hypothesized that the tetramer arrangement has evolved to restrict the flexibility at the active site by buttressing together a pair of dimers. In contrast, DHDPS from methicillin resistant Staphylococcus aureus (MRSA) occurs naturally as a dimer yet retains full activity. Using molecular dynamics simula- tions we have investigated the flexibility of dimeric and tetrameric forms of the E. coli and MRSA enzymes, and reveal that optimal activity is achieved by minimizing the inherent dimer flexibility using two different strategies – by either buttressing two dimers together in the case of the E. coli tetrameric enzyme or strengthening and extending the dimer interface in the dimeric MRSA.

 



Movie 1: E. coli wild-type tetramer dynamics.

 


Movie 2: E. coli L197Y mutant dimer dynamics..

 


Movie 3: Active site dynamics of E. coli tetramer.

 


Movie 4: Active site dynamics of E. coli dimer.

 


Movie 5: MRSA wild-type dimer dynamics.

 



Movie 6: Active site dynamics of MRSA dimer.

 

 

 

 


Conformational properties of the disease-causing Z variant of alpha1 antitrypsin revealed by molecular dynamics simulations

Conformational properties of the disease-causing Z variant of alpha1 antitrypsin revealed by theory and experiment.  Kass, Knaupp, Bottomley, Buckle. (2012).  Biophys. J.102(12):2856-65.

Our investigations identify specific interactions in the breach region of the Z-variant of α1-AT that may play a role in its greater propensity to misfold, aggregate, and thus cause disease, and are consistent with biophysical and structural data that highlight the importance of this region in serpin folding and polymerization.

 


Movie 1. 300-ns Representative Trajectory of Wild-Type α1-AT in Solution.

 


Movie 2. 300-ns Representative Trajectory of Z α1-AT in Solution.

 


Movie 3. 300-ns Representative Trajectory of E342R α1-AT in Solution.

 

 

 

 


Determinants of the substrate specificity of the complement initiating proteases

Wijeyewickrema LC, Yongqing T, Tran TP, Thompson PE, Viljoen JE, Theresa H. Coetzer TH, Duncan RC, Kass I, Buckle AM and Pike RN (2013) Molecular determinants of the substrate specificity of the complement initiating protease, C1r. Journal of Biological Chemistry 288, 15571-15580. doi: 10.1074/jbc.M113.451757

Complement activation represents a crucial innate defence mechanism against invading microorganisms, providing an immediate response against microbial invasion. C1r is responsible for the first enzymatic events in the classical pathway of complement activation. The complement system is strongly implicated in many inflammatory disease states and therefore inhibitors of the initiating proteases could be powerful anti-inflammatory agents.

The data obtained in this study indicates that the specificity of C1r is driven largely by the interaction of the active site of the enzyme with sequences at the cleavage sites of substrates. Examination of the active site specificity of C1r using phage library technology revealed clear specificity for Gln at P2, which are found in these positions in physiological substrates of C1r. Molecular dynamics simulations and structural modeling of the interaction of the C1s activation peptide with the active site of C1r revealed the molecular mechanisms that particularly underpin the specificity of the enzyme for the P2 Gln residue.


Movie 1. A movie of a 100 ns representative MD trajectory of EEKQRIILG bound to C1r protease in solution.

 


Movie 2. A movie of a 100 ns representative MD trajectory of EEKNRIILG bound to C1r protease in solution.