Protein Domain-Swapping Can Be a Consequence of Functional Residues

Abstract

Monomer topology has been implicated in domain-swapping, a potential first step on the route to disease-causing protein aggregation. Despite having the same topology (β1-α1-β2-β3-β4-β5), the cysteine protease inhibitor stefin-B domain swaps more readily than a single-chain variant of the heterodimeric sweet protein monellin (scMn). Here, we computationally study the folding of stefin-B and scMn in order to understand the molecular basis for the difference in their domain-swapping propensities. In agreement with experiments, our structure-based simulations show that scMn folds cooperatively without the population of an intermediate while stefin-B populates an equilibrium intermediate state. Since the simulation intermediate has only one domain structured (β3-β4-β5), it can directly lead to domain-swapping. Using computational variants of stefin-B, we show that the population of this intermediate is caused by regions of stefin-B that have been implicated in protease inhibition. We also find that the protease-binding regions are located on two structural elements and localized in space. In contrast, the residues that contribute to the sweetness of monellin are not localized to a few structural elements but are distributed over the protein fold. We conclude that the distributed functional residues of monellin do not induce large local perturbations in the protein structure, eliminating the formation of folding intermediates and in turn domain-swapping. On the other hand, the localized protease-binding regions of stefin-B promote the formation of a folding intermediate which can lead to domain-swapping. Thus, domain-swapping can be a direct consequence of the constraints that function imposes on the protein structure.

Solubility and Aggregation of Selected Proteins Interpreted on the Basis of Hydrophobicity Distribution

“…The reaction can cause a domino effect. The domain-swapping phenomenon is observed relatively often [ 43 , 44 , 45 ]. The answer to the question about the mechanism of this phenomenon, based on the fuzzy oil drop model, shows that the structures of the hydrophobic core complement one another.…”

New Disulphide Bond in Cystatin-Based Protein Scaffold Prevents Domain-Swap-Mediated Oligomerization and Stabilizes the Functionally Active Form

“…Domain-swap dimerization in the cystatin family has been previously reported to occur through extension of the conserved hydrophobic five-residue “cystatin motif” (QVVAG) in Loop 1 as a consequence of frustration of this hairpin hinge region. 25,26 It has been shown recently that this motif, when engineered into the hinge of a β-hairpin, causes domain swapping of otherwise nondomain-swapped proteins. 27 On the other hand, it has been established that mutations in this hinge region can slow down or even completely eliminate domain-swap oligomerization of cystatins.…”

Human Cellular Retinol Binding Protein II Forms a Domain‐Swapped Trimer Representing a Novel Fold and a New Template for Protein Engineering

“…Providing larger binding surfaces, enhancing the number of binding sites for specific ligands, generating a higher local concentration of active sites, and creating new opportunities for allosteric regulation are some examples of functional diversification through domain‐swapping [6,11–16] . Domain swapping also is a useful protein engineering tool for creating new protein conformational switches, optogenetic tools, artificial enzymes, biosensors and so on as shown by many research groups including us [17–31] . Now our work in engineering the DS trimer as a novel protein engineering template can also be added to this literature.…”

“…[6,[11][12][13][14][15][16] Domain swapping also is a useful protein engineering tool for creating new protein conformational switches, optogenetic tools, artificial enzymes, biosensors and so on as shown by many research groups including us. [17][18][19][20][21][22][23][24][25][26][27][28][29][30][31] Now our work in engineering the DS trimer as a novel protein engineering template can also be added to this literature.…”