Research
My
current research work is to
develope generic and simple polymer models such as Gaussian chains with
Go-model of protein folding to study main-chain conformational changes
in
proteins. The main focus of my research is to understand the influence
of intrinsic plasticity on the mechanism of large-scale conformational
changes (functional transition) and kinetics of flexible proteins.
Complex conformational transitions, induced by ligand binding for example, are often essential to proteins participating in regulatory networks or enzyme catalysis. A protein’s ability to sample a variety of conformational sub-states implies that proteins have an intrinsic flexibility and mobility that influences their function. While experimental measurement can offer direct dynamical information about specific residues, uncovering the detailed mechanisms controlling conformational transitions between two meta-stable states are often elusive. More specifically, experimental studies using nuclear magnetic resonance (NMR) and X-ray provide only a few local structural changes of protein and have not been able to capture the molecular details necessary to fully understand the mechanism of conformational transitions. Whereas atomistic simulations can potentially bridge the gap on time scale up to microsecond, this time scale falls orders of magnitude short for slow protein dynamics (millisecond to second). Also, the use of atomistic approaches becomes computationally inefficient with the increased size of a system. To overcome the problems associated with all-atom simulations, many studies have demonstrated the use of low-resolution (coarse-grained) protein models with simplified representations, such as, only alpha-carbons as point masses and simplified energy functions. Such models require much less computational cost, making them practical to describe the conformational transitions of even large proteins.
To study protein conformational transition mechanism, we have developed a coarse-grained analytical model. Our approach is inspired by a variational model previously developed to characterize protein folding [PRL, 81, 5237, 1998]. The present model accommodates two meta-stable folded conformations coupled by a uniform interpolation parameter as minima of the calculated free energy surface. The natural order parameters of this model are very well suited to describe partially ordered ensembles essential to the conformational dynamics of flexible protein like calmodulin (CaM) and may be an ideal model system to illustrate how conformational flexibility and mobility are a key determinant of its diverse biological functions.
Summary of accomplished work:
Inherent flexibility and protein function
Calcium binding in each domain of CaM induces a “closed” (apo) to “open” (holo) conformational transition that exposes a large hydrophobic surface responsible for molecular recognition of various cellular targets such as myosin light chain kinase. Our studies revealed that inherent flexibility play a crucial role in the apo/holo conformational transition mechanism of the N-terminal CaM domain (nCaM). The resulting detailed transition route from our model is largely consistent with the recently proposed scaffold mechanism in the binding loops of EF-hand family proteins. We find that the N-terminal parts of the calcium binding loops shows higher flexibility than the C-terminal parts which form this scaffold structure. Our model predicts that binding loop II, with higher flexibility and earlier structural change than binding loop I, dominates the open/closed conformational transition in nCaM.
Conformational transition by cracking
We explore how inherent flexibility of a protein molecule influences the mechanism controlling allosteric transitions. The striking differences in the predicted transition mechanism for the opening of the two domains of CaM emphasize that inherent flexibility is the key determinant of the complex conformational changes that occur in proteins. In particular, the C-terminal domain of CaM (cCaM), which is inherently less flexible than nCaM, reveals cracking or local partial unfolding during the open/closed transition. This result is in harmony with the picture that cracking relieves local stresses of sufficiently rigid regions of protein caused by conformational deformations. We also compare the conformational transition in a recently studied even-odd paired fragment of CaM. Our results rationalize the different relative binding affinities of the EF-hands in the engineered fragment compared with the intact odd-even paired EF-hands (nCaM and cCaM) in terms of changes in flexibility along the transition route. Aside from elucidating general theoretical ideas about the cracking mechanism, these studies also emphasize how the remarkable intrinsic plasticity of CaM underlies conformational dynamics essential for its diverse functions.
Interplay among topology, plasticity and energetics
We investigate the interplay between topology, structural plasticity and energetics of the proteins from their functional transitions. Recently, we found that the open/closed conformational transition mechanism of the two topologically similar and homologous CaM domains are distinctly different due to cracking in cCaM. To analyze cracking in more detail we calculate strain energy for each residue of the CaM domains from their deformed structures. This result agrees very well with the change in conformational flexibility of the corresponding CaM domains along their transition pathway. We also found that residues of the cCaM which crack release local stresses. Furthermore, our study elucidates that cracking results in lower free energy barriers and more efficient activity. The above findings confirm that functional transitions in proteins are predetermined by their intrinsic plasticity encoded in the amino acid sequence network (contact map).
Summary of ongoing work:
Ongoing research is focused to study conformational transitions in other protein systems such as inactive/active interconversion of nitrogen regulatory protein C (NtrC), a member of the phosphorylation-mediated signaling family. We are also investigating the complex inter-relationship between stability and conformational flexibility by characterizing the ensemble of intermediate conformational states of protein systems. This is important as a proper balance between stability and flexibility of protein is essential for its diverse biological functions. The final goal is to extend our present model to study protein-ligand binding interactions. Our initial attempt is to explore protein-ligand interactions for ligands such as metal ions and small molecules. We are mainly focusing to understand the influence of conformational flexibility of proteins on the binding affinity and the kinetics of binding.
Complex conformational transitions, induced by ligand binding for example, are often essential to proteins participating in regulatory networks or enzyme catalysis. A protein’s ability to sample a variety of conformational sub-states implies that proteins have an intrinsic flexibility and mobility that influences their function. While experimental measurement can offer direct dynamical information about specific residues, uncovering the detailed mechanisms controlling conformational transitions between two meta-stable states are often elusive. More specifically, experimental studies using nuclear magnetic resonance (NMR) and X-ray provide only a few local structural changes of protein and have not been able to capture the molecular details necessary to fully understand the mechanism of conformational transitions. Whereas atomistic simulations can potentially bridge the gap on time scale up to microsecond, this time scale falls orders of magnitude short for slow protein dynamics (millisecond to second). Also, the use of atomistic approaches becomes computationally inefficient with the increased size of a system. To overcome the problems associated with all-atom simulations, many studies have demonstrated the use of low-resolution (coarse-grained) protein models with simplified representations, such as, only alpha-carbons as point masses and simplified energy functions. Such models require much less computational cost, making them practical to describe the conformational transitions of even large proteins.
To study protein conformational transition mechanism, we have developed a coarse-grained analytical model. Our approach is inspired by a variational model previously developed to characterize protein folding [PRL, 81, 5237, 1998]. The present model accommodates two meta-stable folded conformations coupled by a uniform interpolation parameter as minima of the calculated free energy surface. The natural order parameters of this model are very well suited to describe partially ordered ensembles essential to the conformational dynamics of flexible protein like calmodulin (CaM) and may be an ideal model system to illustrate how conformational flexibility and mobility are a key determinant of its diverse biological functions.
Summary of accomplished work:
Inherent flexibility and protein function
Calcium binding in each domain of CaM induces a “closed” (apo) to “open” (holo) conformational transition that exposes a large hydrophobic surface responsible for molecular recognition of various cellular targets such as myosin light chain kinase. Our studies revealed that inherent flexibility play a crucial role in the apo/holo conformational transition mechanism of the N-terminal CaM domain (nCaM). The resulting detailed transition route from our model is largely consistent with the recently proposed scaffold mechanism in the binding loops of EF-hand family proteins. We find that the N-terminal parts of the calcium binding loops shows higher flexibility than the C-terminal parts which form this scaffold structure. Our model predicts that binding loop II, with higher flexibility and earlier structural change than binding loop I, dominates the open/closed conformational transition in nCaM.
Conformational transition by cracking
We explore how inherent flexibility of a protein molecule influences the mechanism controlling allosteric transitions. The striking differences in the predicted transition mechanism for the opening of the two domains of CaM emphasize that inherent flexibility is the key determinant of the complex conformational changes that occur in proteins. In particular, the C-terminal domain of CaM (cCaM), which is inherently less flexible than nCaM, reveals cracking or local partial unfolding during the open/closed transition. This result is in harmony with the picture that cracking relieves local stresses of sufficiently rigid regions of protein caused by conformational deformations. We also compare the conformational transition in a recently studied even-odd paired fragment of CaM. Our results rationalize the different relative binding affinities of the EF-hands in the engineered fragment compared with the intact odd-even paired EF-hands (nCaM and cCaM) in terms of changes in flexibility along the transition route. Aside from elucidating general theoretical ideas about the cracking mechanism, these studies also emphasize how the remarkable intrinsic plasticity of CaM underlies conformational dynamics essential for its diverse functions.
Interplay among topology, plasticity and energetics
We investigate the interplay between topology, structural plasticity and energetics of the proteins from their functional transitions. Recently, we found that the open/closed conformational transition mechanism of the two topologically similar and homologous CaM domains are distinctly different due to cracking in cCaM. To analyze cracking in more detail we calculate strain energy for each residue of the CaM domains from their deformed structures. This result agrees very well with the change in conformational flexibility of the corresponding CaM domains along their transition pathway. We also found that residues of the cCaM which crack release local stresses. Furthermore, our study elucidates that cracking results in lower free energy barriers and more efficient activity. The above findings confirm that functional transitions in proteins are predetermined by their intrinsic plasticity encoded in the amino acid sequence network (contact map).
Summary of ongoing work:
Ongoing research is focused to study conformational transitions in other protein systems such as inactive/active interconversion of nitrogen regulatory protein C (NtrC), a member of the phosphorylation-mediated signaling family. We are also investigating the complex inter-relationship between stability and conformational flexibility by characterizing the ensemble of intermediate conformational states of protein systems. This is important as a proper balance between stability and flexibility of protein is essential for its diverse biological functions. The final goal is to extend our present model to study protein-ligand binding interactions. Our initial attempt is to explore protein-ligand interactions for ligands such as metal ions and small molecules. We are mainly focusing to understand the influence of conformational flexibility of proteins on the binding affinity and the kinetics of binding.