The electrostatic profile of consecutive Cβ atoms applied to protein structure quality assessment.

The structure of a protein provides insight into its physiological interactions with other components of the cellular soup. Methods that predict putative structures from sequences typically yield multiple, closely-ranked possibilities. A critical component in the process is the model quality assessing program (MQAP), which selects the best candidate from this pool of structures. Here, we present a novel MQAP based on the physical properties of sidechain atoms. We propose a method for assessing the quality of protein structures based on the electrostatic potential difference (EPD) of Cβ atoms in consecutive residues. We demonstrate that the EPDs of Cβ atoms on consecutive residues provide unique signatures of the amino acid types. The EPD of Cβ atoms are learnt from a set of 1000 non-homologous protein structures with a resolution cuto of 1.6 Å obtained from the PISCES database. Based on the Boltzmann hypothesis that lower energy conformations are proportionately sampled more, and on Annsen's thermodynamic hypothesis that the native structure of a protein is the minimum free energy state, we hypothesize that the deviation of observed EPD values from the mean values obtained in the learning phase is minimized in the native structure. We achieved an average specificity of 0.91, 0.94 and 0.93 on hg_structal, 4state_reduced and ig_structal decoy sets, respectively, taken from the Decoys `R' Us database. The source code and manual is made available at https://github.com/sanchak/mqap and permanently available on 10.5281/zenodo.7134

Characterizing alpha helical properties of Ebola viral proteins as potential targets for inhibition of alpha-helix mediated protein-protein interactions

Ebola, considered till recently as a rare and endemic disease, has dramatically transformed into a potentially global humanitarian crisis. The genome of Ebola, a member of the Filoviridae family, encodes seven proteins. Based on the recently implemented software (PAGAL) for analyzing the hydrophobicity and amphipathicity properties of alpha helices (AH) in proteins, we characterize the helices in the Ebola proteome. We demonstrate that AHs with characteristically unique features are involved in critical interactions with the host proteins. For example, the Ebola virus membrane fusion subunit, GP2, from the envelope glycoprotein ectodomain has an AH with a large hydrophobic moment. The neutralizing antibody (KZ52) derived from a human survivor of the 1995 Kikwit outbreak recognizes a protein epitope on this AH, emphasizing the critical nature of this secondary structure in the virulence of the Ebola virus. Our method ensures a comprehensive list of such `hotspots'. These helices probably are or can be the target of molecules designed to inhibit AH mediated protein-protein interactions. Further, by comparing the AHs in proteins of the related Marburg viruses, we are able to elicit subtle changes in the proteins that might render them ineffective to previously successful drugs. Such differences are difficult to identify by a simple sequence or structural alignment. Thus, analyzing AHs in the small Ebola proteome can aid rational design aimed at countering the `largest Ebola epidemic, affecting multiple countries in West Africa

Protein structure quality assessment based on the distance profiles of consecutive backbone Cα atoms

Predicting the three dimensional native state structure of a protein from its primary sequence is an unsolved grand challenge in molecular biology. Two main computational approaches have evolved to obtain the structure from the protein sequence - ab initio/de novo methods and template-based modeling - both of which typically generate multiple possible native state structures. Model quality assessment programs (MQAP) validate these predicted structures in order to identify the correct native state structure. Here, we propose a MQAP for assessing the quality of protein structures based on the distances of consecutive Cα atoms. We hypothesize that the root-mean-square deviation of the distance of consecutive Cα (RDCC) atoms from the ideal value of 3.8 Å, derived from a statistical analysis of high quality protein structures (top100H database), is minimized in native structures. Based on tests with the top100H set, we propose a RDCC cutoff value of 0.012 Å, above which a structure can be filtered out as a non-native structure. We applied the RDCC discriminator on decoy sets from the Decoys 'R' Us database to show that the native structures in all decoy sets tested have RDCC below the 0.012 Å cutoff. While most decoy sets were either indistinguishable using this discriminator or had very few violations, all the decoy structures in the fisa decoy set were discriminated by applying the RDCC criterion. This highlights the physical non-viability of the fisa decoy set, and possible issues in benchmarking other methods using this set. The source code and manual is made available at https://github.com/sanchak/mqap and permanently available on 10.5281/zenodo.7134

The electrostatic profile of consecutive Cβ atoms applied to protein structure quality assessment

The structure of a protein provides insight into its physiological interactions with other components of the cellular soup. Methods that predict putative structures from sequences typically yield multiple, closely-ranked possibilities. A critical component in the process is the model quality assessing program (MQAP), which selects the best candidate from this pool of structures. Here, we present a novel MQAP based on the physical properties of sidechain atoms. We propose a method for assessing the quality of protein structures based on the electrostatic potential difference (EPD) of Cβ atoms in consecutive residues. We demonstrate that the EPDs of Cβ atoms on consecutive residues provide unique signatures of the amino acid types. The EPD of Cβ atoms are learnt from a set of 1000 non-homologous protein structures with a resolution cuto of 1.6 Å obtained from the PISCES database. Based on the Boltzmann hypothesis that lower energy conformations are proportionately sampled more, and on Annsen's thermodynamic hypothesis that the native structure of a protein is the minimum free energy state, we hypothesize that the deviation of observed EPD values from the mean values obtained in the learning phase is minimized in the native structure. We achieved an average specificity of 0.91, 0.94 and 0.93 on hg_structal, 4state_reduced and ig_structal decoy sets, respectively, taken from the Decoys 'R' Us database. The source code and manual is made available at https://github.com/sanchak/mqap and permanently available on 10.5281/zenodo.7134

Directed evolution induces tributyrin hydrolysis in a virulence factor of Xylella fastidiosa using a duplicated gene as a template

Duplication of genes is one of the preferred ways for natural selection to add advantageous functionality to the genome without having to reinvent the wheel with respect to catalytic efficiency and protein stability. The duplicated secretory virulence factors of Xylella fastidiosa (LesA, LesB and LesC), implicated in Pierce's disease of grape and citrus variegated chlorosis of citrus species, epitomizes the positive selection pressures exerted on advantageous genes in such pathogens. A deeper insight into the evolution of these lipases/esterases is essential to develop resistance mechanisms in transgenic plants. Directed evolution, an attempt to accelerate the evolutionary steps in the laboratory, is inherently simple when targeted for loss of function. A bigger challenge is to specify mutations that endow a new function, such as a lost functionality in a duplicated gene. Previously, we have proposed a method for enumerating candidates for mutations intended to transfer the functionality of one protein into another related protein based on the spatial and electrostatic properties of the active site residues (DECAAF). In the current work, we present in vivo validation of DECAAF by inducing tributyrin hydrolysis in LesB based on the active site similarity to LesA. The structures of these proteins have been modeled using RaptorX based on the closely related LipA protein from Xanthomonas oryzae. These mutations replicate the spatial and electrostatic conformation of LesA in the modeled structure of the mutant LesB as well, providing in silico validation before proceeding to the laborious in vivo work. Such focused mutations allows one to dissect the relevance of the duplicated genes in finer detail as compared to gene knockouts, since they do not interfere with other moonlighting functions, protein expression levels or protein-protein interaction

The PDB database is a rich source of alpha-helical anti-microbial peptides to combat disease causing pathogens

The therapeutic potential of α-helical anti-microbial peptides (AH-AMP) to combat pathogens is fast gaining prominence. Based on recently published open access software for characterizing α-helical peptides (PAGAL), we elucidate a search methodology (SCALPEL) that leverages the massive structural data pre-existing in the PDB database to obtain AH-AMPs belonging to the host proteome. We provide in vitro validation of SCALPEL on plant pathogens (Xylella fastidiosa, Xanthomonas arboricola and Liberibacter crescens) by identifying AH-AMPs that mirror the function and properties of cecropin B, a well-studied AH-AMP. The identified peptides include a linear AH-AMP present within the existing structure of phosphoenolpyruvate carboxylase (PPC20), and an AH-AMP mimicing the properties of the two α-helices of cecropin B from chitinase (CHITI25). The minimum inhibitory concentration of these peptides are comparable to that of cecropin B, while anionic peptides used as control failed to show any inhibitory effect on these pathogens. Substitute therapies in place of conventional chemotherapies using membrane permeabilizing peptides like these might also prove effective to target cancer cells. The use of native structures from the same organism could possibly ensure that administration of such peptides will be better tolerated and not elicit an adverse immune response. We suggest a similar approach to target Ebola epitopes, enumerated using PAGAL recently, by selecting suitable peptides from the human proteome, especially in wake of recent reports of cationic amphiphiles inhibiting virus entry and infection

Fracture strength of zirconia implant abutments on narrow diameter implants with internal and external implant abutment connections: A study on the titanium resin base concept

BACKGROUND There is limited knowledge regarding the strength of zirconia abutments with internal and external implant abutment connections and zirconia abutments supported by a titanium resin base (Variobase, Straumann) for narrow diameter implants. OBJECTIVES To compare the fracture strength of narrow diameter abutments with different types of implant abutment connections after chewing simulation. MATERIAL AND METHODS Hundred and twenty identical customized abutments with different materials and implant abutment connections were fabricated for five groups: 1-piece zirconia abutment with internal connection (T1, Cares-abutment-Straumann BL-NC implant, Straumann Switzerland), 1-piece zirconia abutment with external hex connection (T2, Procera abutment-Branemark NP implant, Nobel Biocare, Sweden), 2-piece zirconia abutments with metallic insert for internal connection (T3, Procera abutment-Replace NP implant, Nobel Biocare), 2-piece zirconia abutment on titanium resin base (T4, LavaPlus abutment-VarioBase-Straumann BL-NC implant, 3M ESPE, Germany) and 1-piece titanium abutment with internal connection (C, Cares-abutment-Straumann BL-NC implant, Straumann, Switzerland). All implants had a narrow diameter ranging from 3.3 to 3.5 mm. Sixty un-restored abutments and 60 abutments restored with glass-ceramic crowns were tested. Mean bending moments were compared using ANOVA with p-values adjusted for multiple comparisons using Tukey's procedure. RESULTS The mean bending moments were 521 ± 33 Ncm (T4), 404 ± 36 Ncm (C), 311 ± 106 Ncm (T1) 265 ± 22 Ncm (T3) and 225 ± 29 (T2) for un-restored abutments and 278 ± 84 Ncm (T4), 302 ± 170 Ncm (C), 190 ± 55 Ncm (T1) 80 ± 102 Ncm (T3) and 125 ± 57 (T2) for restored abutments. For un-restored abutments, C and T4 had similar mean bending moments, significantly higher than those of the three other groups (p < .05). Titanium abutments (C) had significantly higher bending moments than identical zirconia abutments (T1) (p < .05). Zirconia abutments (T1) with internal connection had higher bending moments than zirconia abutments with external connection (T2) (p < .05). For all test groups, the bending moments were significantly reduced when restored with all-ceramic crowns. CONCLUSIONS For narrow diameter abutments, the fracture strength of 2-piece internal connected zirconia abutments fixed on titanium resin bases was similar to those obtained for 1-piece titanium abutments. Narrow diameter zirconia abutments with internal connection exhibited higher fracture strength than zirconia abutments with an external connection. Titanium abutments with an internal connection were significantly stronger than identical zirconia abutments