Conformational fluctuations versus constraints in amino acid side chains: the evolution of information content from free amino acids to proteins

Like all other complex biological systems, proteins exhibit properties not found in free amino acids (i.e., emergent properties). Here, we explore top-down constraints experienced by the residue side chains in proteins compared to amino acids in increasingly complex molecular environments: free amino acids, end-capped amino acids, and the central residue in an alpha-helical nonapeptide. The crystalline structure of the contractile protein profilin Ib and the enzyme trypsin were chosen as objects of study, and submitted to 10 ns molecular dynamics (MD) simulations. The results revealed increased conformational constraints on the side chains when going from the simpler to the more complex compounds. A Shannon entropy (SE) analysis of the conformational behavior of the side chains showed in most cases a progressive and marked decrease in the SE of the chi1 and chi2 dihedral angles. This is equivalent to stating that conformational constraints on the side chain of residues increase their information content and, hence, recognition specificity compared to free amino acids. In other words, the vastly increased information content of a protein relative to its free monomers is embedded not only in the tertiary structure of the backbone, but also in the conformational behavior of the side chains. The postulated implication is that both backbone and side chains, by virtue of being conformationally constrained, contribute to the protein's recognition specificity toward other macromolecules and ligands.     

Immunoglobulin kappa light chain and its amyloidogenic mutants: a molecular dynamics study

AL amyloidosis and LCDD are pathological conditions caused by extracellural deposition of monoclonal Ig light chain variable domains. In the former case, deposits have a form of amyloid fibrils, in the latter, amorphous aggregates. 1REI kappa light chain variable domain and its two point mutants, R61N and D82I, were chosen for the analysis in this work. Wild 1REI does not create deposits in vitro, while R61N aggregates as amyloid fibrils and D82I creates amorphous aggregates. Both mutated residues create a conserved salt bridge; thus, substitution of any of them should decrease V(L) domain stability. For these three proteins, 5 ns MD simulations were conducted in temperatures of 300 K and 400 K, with protonated and unprotonated acidic residues, mimicking acidic and neutral experimental pH conditions (3 sets: N300, N400, and A400). The analysis of trajectories focused on characterization of changes in conformational behavior and stability of Ig kappa light chain variable domain caused by single aminoacid substitutions that were experimentally proved to enhance aggregation propensity, both in the form of amyloid and amorphous aggregates. Residue D82 turns out to be involved not only in R61-D82 but also in K45-D82 interaction, which was not observed in the X-ray structure, but frequently populated simulations of 1REI. The substitution D82I excludes both interactions, resulting in substantial destabilization (i.e., easier aggregation). Examination of behavior of edge regions of V(L) beta-sandwich reveals significant alterations in D82I mutant compared to wild 1REI, while relatively small changes occur in R61N. This suggests that mild and slow destabilization is the reason of the conversion of V(L) to partially folded amyloidogenic intermediate structure.     

Reconstitution of the type-1 active site of the H145G/A variants of nitrite reductase by ligand insertion

Variants of the copper-containing nitrite reductase (NiR) of Alcaligenes faecalis S6 were constructed by site-directed mutagenesis, by which the C-terminal histidine ligand (His145) of the Cu in the type-1 site was replaced by an alanine or a glycine. The type-1 sites in the NiR variants as isolated, are in the reduced form, but can be oxidized in the presence of external ligands, like (substituted) imidazoles and chloride. The reduction potential of the type-1 site of NiR-H145A reconstituted with imidazole amounts to 505 mV vs NHE (20 degrees C, pH 7, 10 mM imidazole), while for the native type-1 site it amounts to 260 mV. XRD data on crystals of the reduced and oxidized NiR-H145A variant show that in the reduced type-1 site the metal is 3-coordinated, but in the oxidized form takes up a ligand from the solution. With the fourth (exogenous) ligand in place the type-1 site is able to accept electrons at about the same rate as the wt NiR, but it is unable to pass the electron onto the type-2 site, leading to loss of enzymatic activity. It is argued that the uptake of an electron by the mutated type-1 site is accompanied by a loss of the exogenous ligand and a concomitant rise of the redox potential. This rise effectively traps the electron in the type-1 site.     

Internal force field in proteins seen by divergence entropy

The characteristic distribution of non-binding interactions in a protein is described. It establishes that hydrophobic interactions can be characterized by suitable 3D Gauss functions while electrostatic interactions generally follow a random distribution. The implementation of this observation suggests differentiated optimization procedure for these two types of interactions. The electrostatic interaction may follow traditional energy optimization while the criteria for convergence shall measure the accordance with 3-D Gauss function.