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Bjørn O Brandsdal

Publications and source records attributed to Bjørn O Brandsdal.

10 recordsLinked to original sources
PubMed2006

Optimization of electrostatics as a strategy for cold-adaptation: a case study of cold- and warm-active elastases.

Adaptation to both high and low temperatures requires proteins with special properties. While organisms living at or close to the boiling point of water need to have proteins with increased stability, other properties are required at temperatures close to the freezing point of water. Indeed, it has been shown that enzymes adapted to cold environments are less resistant to heat with a concomitant increased activity as compared to their warm-active counter-parts. Several recent studies have pointed in the direction that electrostatic interactions play a central role in temperature adaptation, and in this study we investigate the role such interactions have in adaptation of elastase from Atlantic salmon and pig. Molecular dynamics (MD) simulations have been used to generate structural ensembles at 283 and 310 K of the psychrophilic and mesophilic elastase, and a total of eight 12 ns simulations have been carried out. Even though the two homologues have a highly similar three-dimensional structure, the location and number of charged amino acids are very different. Based on the simulated structures we find that very few salt-bridges are stable throughout the simulations, and provide little stabilization/destabilization of the proteins as judged by continuum electrostatic calculations. However, the mesophilic elastase is characterized by a greater number of salt-bridges as well as a putative salt-bridge network close to the catalytic site, indicating a higher rigidity of the components involved in the catalytic cycle. In addition, subtle differences are also found in the electrostatic potentials in the vicinity of the catalytic residues, which may explain the increased catalytic efficiency of the cold-adapted elastase.

Acclimatization↗
PubMed2006

Free energy calculations show that acidic P1 variants undergo large pKa shifts upon binding to trypsin.

Serine proteinases and their protein inhibitors belong to one of the most comprehensively studied models of protein-protein interactions. It is well established that the narrow trypsin specificity is caused by the presence of a negatively charged aspartate at the specificity pocket. X-ray crystallography as well as association measurements revealed, surprisingly, that BPTI with glutamatic acid as the primary binding (P1) residue was able to bind to trypsin. Previous free energy calculations showed that there was a substantially unfavorable binding free energy associated with accommodation of ionized P1 Glu at the S1-site of trypsin. In this study, the binding of P1 Glu to trypsin has been systematically investigated in terms of the protonation states of P1 Glu and Asp189, the orientation of Gln192, as well as the possible presence of counterions using the linear interaction energy (LIE) approach and the free energy perturbation (FEP) method. Twenty-four conceivable binding arrangements were evaluated and quantitative agreement with experiments is obtained when the P1 Glu binds in its protonated from. The results suggest that P1 Glu is one of the variants of BPTI that inhibit trypsin strongest at low pH, contrary to the specificity profile of trypsin, suggesting a new regulation mechanism of trypsin-like enzymes.

Aprotinin↗
PubMed2006

Predicting proteinase specificities from free energy calculations.

The role of the primary binding residue (P1) in complexes between three different subtilases (subtilisin Carlsberg, thermitase and proteinase K) and their canonical protein inhibitor eglin c have been studied by free energy calculations. Based on the crystal structures of eglin c in complex with subtilisin Carlsberg and thermitase, and a homology model of the eglin c-proteinase K complex, a total of 57 mutants have been constructed and docked into their host proteins. The binding free energy was then calculated using molecular dynamics (MD) simulations combined with the linear interaction energy (LIE) method for all complexes differing only in the nature of the amino acid at the P1 position. LIE calculations for 19 different complexes for each subtilase were thus carried out excluding proline. The effects of substitutions at the P1 position on the binding free energies are found to be very large, and positively charged residues (Arg, Lys and His) are particularly deleterious for all three enzymes. The charged variants of the acidic side chains are found to bind more favorably as compared to their protonated states in all three subtilases. Furthermore, hydrophobic amino acids are accommodated most favorably at the S1-site in all three enzymes. Comparison of the three series of binding free energies shows only minor differences in the 19 computed relative binding free energies among these subtilases. This is further reflected in the correlation coefficient between the 23 relative binding free energies obtained, including the possible protonation states of ionizable side chains, but excluding the P1 Pro, for subtilisin Carlsberg versus thermitase (0.95), subtilisin versus proteinase K (0.94) and thermitase versus proteinase K (0.96).

Algorithms↗
PubMed2005

Probing the effect of point mutations at protein-protein interfaces with free energy calculations.

We have studied the effect of point mutations of the primary binding residue (P1) at the protein-protein interface in complexes of chymotrypsin and elastase with the third domain of the turkey ovomucoid inhibitor and in trypsin with the bovine pancreatic trypsin inhibitor, using molecular dynamics simulations combined with the linear interaction energy (LIE) approach. A total of 56 mutants have been constructed and docked into their host proteins. The free energy of binding could be reliably calculated for 52 of these mutants that could unambiguously be fitted into the binding sites. We find that the predicted binding free energies are in very good agreement with experimental data with mean unsigned errors between 0.50 and 1.03 kcal/mol. It is also evident that the standard LIE model used to study small drug-like ligand binding to proteins is not suitable for protein-protein interactions. Three different LIE models were therefore tested for each of the series of protein-protein complexes included, and the best models for each system turn out to be very similar. The difference in parameterization between small drug-like compounds and protein point mutations is attributed to the preorganization of the binding surface. Our results clearly demonstrate the potential of free energy calculations for probing the effect of point mutations at protein-protein interfaces and for exploring the principles of specificity of hot spots at the interface.

Animals↗
PubMed2005

Resolving the energy paradox of chaperone/usher-mediated fibre assembly.

Periplasmic chaperone/usher machineries are used for assembly of filamentous adhesion organelles of Gram-negative pathogens in a process that has been suggested to be driven by folding energy. Structures of mutant chaperone-subunit complexes revealed a final folding transition (condensation of the subunit hydrophobic core) on the release of organelle subunit from the chaperone-subunit pre-assembly complex and incorporation into the final fibre structure. However, in view of the large interface between chaperone and subunit in the pre-assembly complex and the reported stability of this complex, it is difficult to understand how final folding could release sufficient energy to drive assembly. In the present paper, we show the X-ray structure for a native chaperone-fibre complex that, together with thermodynamic data, shows that the final folding step is indeed an essential component of the assembly process. We show that completion of the hydrophobic core and incorporation into the fibre results in an exceptionally stable module, whereas the chaperone-subunit pre-assembly complex is greatly destabilized by the high-energy conformation of the bound subunit. This difference in stabilities creates a free energy potential that drives fibre formation.

Bacterial Proteins↗
PubMed2005

Increased flexibility as a strategy for cold adaptation: a comparative molecular dynamics study of cold- and warm-active uracil DNA glycosylase.

Uracil DNA glycosylase (UDG) is a DNA repair enzyme in the base excision repair pathway and removes uracil from the DNA strand. Atlantic cod UDG (cUDG), which is a cold-adapted enzyme, has been found to be up to 10 times more catalytically active in the temperature range 15-37 degrees C as compared with the warm-active human counterpart. The increased catalytic activity of cold-adapted enzymes as compared with their mesophilic homologues are partly believed to be caused by an increase in the structural flexibility. However, no direct experimental evidence supports the proposal of increased flexibility of cold-adapted enzymes. We have used molecular dynamics simulations to gain insight into the structural flexibility of UDG. The results from these simulations show that an important loop involved in DNA recognition (the Leu(272) loop) is the most flexible part of the cUDG structure and that the human counterpart has much lower flexibility in the Leu(272) loop. The flexibility in this loop correlates well with the experimental k(cat)/K(m) values. Thus, the data presented here add strong support to the idea that flexibility plays a central role in adaptation to cold environments.

Adaptation, Physiological↗
PubMed2004

Binding affinity prediction with different force fields: examination of the linear interaction energy method.

A systematic study of the linear interaction energy (LIE) method and the possible dependence of its parameterization on the force field and system (receptor binding site) is reported. We have calculated the binding free energy for nine different ligands in complex with P450cam using three different force fields (Amber95, Gromos87, and OPLS-AA). The results from these LIE calculations using our earlier parameterization give relative free energies of binding that agree remarkably well with the experimental data. However, the absolute energies are too positive for all three force fields, and it is clear that an additional constant term (gamma) is required in this case. Out of five examined LIE models, the same one emerges as the best for all three force fields, and this, in fact, corresponds to our earlier one apart from the addition of the constant gamma, which is almost identical for the three force fields. Thus, the present free energy calculations clearly indicate that the coefficients of the LIE method are independent of the force field used. Their relation to solvation free energies is also demonstrated. The only free parameter of the best model is gamma, which is found to depend on the hydrophobicity of the binding site. We also attempt to quantify the binding site hydrophobicity of four different proteins which shows that the ordering of gamma's for these sites reflects the fraction of hydrophobic surface area.

Journal Article↗
PubMed2004

The primary structure and specificity determining residues displayed by recombinant salmon antibody domains.

Previously, single chain fragments of salmon (Salmo salar L.) immunoglobulin variable regions (scFv) were isolated by reactivity towards trinitrophenyl (TNP) or fluorescein (FITC) using phage display technology. The fine specificity of six scFv clones were analysed by ELISA, while the primary structure was determined by DNA sequencing. In addition, preliminary models of one anti-TNP and one anti-FITC clone were built. Here, a follow-up analysis of the primary and tertiary structure of all six clones is focused on the structural basis for hapten specificity. Tertiary structure was analysed by molecular modelling of the antigen combining site. The analysis shows that reactivity to each hapten is maintained by a number of different combinations of VH, D, JH and VL sequences. Accordingly, various sizes of CDR3 on both the heavy and light chain and CDR2 of IgH may support TNP binding. Due to variability of the antigen combining site each clone probably has a distinct binding affinity. However, a feature common among the four scFv antibodies that recognise TNP is a positively charged Arg in CDR2 of either the heavy or light chain. In the majority of the anti-TNP clones localisation of this side-chain is stabilised by a negatively charged Asp in LCDR1. In addition, a Trp in LCDR3 is conserved in all the anti-TNP clones. Also, the anti-FITC clones display a Trp in the LCDR3, suggesting its participation in binding of FITC as well. In combination with a large aromatic amino acid near the N-terminus of HCDR2 and a positively charged Arg in CDR1, these residues probably determine both specificity and affinity towards the FITC moiety.

Amino Acid Sequence↗
PubMed2002

Ligand binding affinities from MD simulations.

Simplified free energy calculations based on force field energy estimates of ligand-receptor interactions and thermal conformational sampling have emerged as a useful tool in structure-based ligand design. Here we give an overview of the linear interaction energy (LIE) method for calculating ligand binding free energies from molecular dynamics simulations. A notable feature is that the binding energetics can be predicted by considering only the intermolecular interactions of the ligand in the associated and dissociated states. The approximations behind this approach are examined, and different parametrizations of the model are discussed. LIE-type methods appear particularly promising for computational "lead optimization". Recent applications to protein-protein interactions and ion channel blocking are also discussed.

Animals↗