Electrostatic Fingerprint Research Articles

Mapping the favourable character of crystal and intermolecular contacts by electrostatic fingerprints

Mapping the favourable character of crystal and intermolecular contacts by electrostatic fingerprints

Comparative Surface Electrostatics and Normal Mode Analysis of High and Low Pathogenic H7N7 Avian Influenza Viruses.

Influenza A viruses are rarely symptomatic in wild birds, while representing a higher threat to poultry and mammals, where they can cause a variety of symptoms, including death. H5 and H7 subtypes of influenza viruses are of particular interest because of their pathogenic potential and reported capacity to spread from poultry to mammals, including humans. The identification of molecular fingerprints for pathogenicity can help surveillance and early warning systems, which are crucial to prevention and protection from such potentially pandemic agents. In the past decade, comparative analysis of the surface features of hemagglutinin, the main protein antigen in influenza viruses, identified electrostatic fingerprints in the evolution and spreading of H5 and H9 subtypes. Electrostatic variation among viruses from avian or mammalian hosts was also associated with host jump. Recent findings of fingerprints associated with low and highly pathogenic H5N1 viruses, obtained by means of comparative electrostatics and normal modes analysis, prompted us to check whether such fingerprints can also be found in the H7 subtype. Indeed, evidence presented in this work showed that also in H7N7, hemagglutinin proteins from low and highly pathogenic strains present differences in surface electrostatics, while no meaningful variation was found in normal modes.

Electrostatic Fingerprints of Catalytically Active Amino Acids in Enzymes

The computed electrostatic and proton transfer properties are studied for 20 enzymes that represent all six major EC classes and a variety of different folds. The properties of aspartate, glutamate, and lysine residues that have been previously experimentally determined to be catalytically active are reported. All catalytic aspartate and glutamate residues studied here are strongly coupled to at least one other aspartate or glutamate residue and often to multiple other carboxylate residues with intrinsic pKa differences less than 1 pH unit. Sometimes these catalytic acidic residues are also coupled to a histidine residue, such that the intrinsic pKa of the acidic residue is higher than that of the histidine. All catalytic lysine residues studied here are strongly coupled to tyrosine or cysteine residues, wherein the intrinsic pKa of the anion‐forming residue is higher than that of the lysine. Some catalytic lysines are also coupled to other lysines with intrinsic pKa differences within 1 pH unit. Some evidence of the possible types of interactions that facilitate nucleophilicity is discussed. The interactions reported here provide important clues about how side chain functional groups that are weak Br⊘nsted acids or bases for the free amino acid in solution can become strong acids, bases or nucleophiles in the enzymatic environment. Such interactions are important considerations in enzyme design.

Electrostatic fingerprints of catalytically active amino acids in enzymes.

The computed electrostatic and proton transfer properties are studied for 20 enzymes that represent all six major enzyme commission classes and a variety of different folds. The properties of aspartate, glutamate, and lysine residues that have been previously experimentally determined to be catalytically active are reported. The catalytic aspartate and glutamate residues studied here are strongly coupled to at least one other aspartate or glutamate residue and often to multiple other carboxylate residues with intrinsic pKa differences less than 1 pH unit. Sometimes these catalytic acidic residues are also coupled to a histidine residue, such that the intrinsic pKa of the acidic residue is higher than that of the histidine. All catalytic lysine residues studied here are strongly coupled to tyrosine or cysteine residues, wherein the intrinsic pKa of the anion-forming residue is higher than that of the lysine. Some catalytic lysines are also coupled to other lysines with intrinsic pKa differences within 1 pH unit. Some evidence of the possible types of interactions that facilitate nucleophilicity is discussed. The interactions reported here provide important clues about how side chain functional groups that are weak Brønsted acids or bases for the free amino acid in solution can achieve catalytic potency and become strong acids, bases or nucleophiles in the enzymatic environment.

Antimicrobial effects of syndiotactic polypeptides

We present design and antibacterial studies of stereochemically diversified antimicrobial peptides against multidrug-resistant bacterial pathogens. Syndiotactic polypeptides are polymers of alternating L and D amino acids with LDLD or DLDL backbone stereochemical sequence, which can form stable gramicidin like helical conformations. We designed, synthesized and characterized eight model molecular systems with varied electrostatic fingerprints, modulated through calibrated sequence positioning. Six out of eight model systems showed very impressive antimicrobial activity against three difficult to treat bacterial species, Gentamicin resistant MRSA, E. coli and Mycobacterium. More importantly, the designed LDLD peptides were equally potent in serum, an important drawback of poly L peptide sequences due to enzyme mediated degradation and ion sensitivity. Further, we tested the activity of the designed peptides against drug-resistant clinical isolates of Staphylococcus aureus and Escherichia coli. Molecular dynamics simulation studies suggest formation of an assembly of individual peptides, preceding the membrane interaction and deformation. The activity estimates are comparable with the available peptide based antimicrobials, and are also highly specific and less toxic as per standard estimates. Incorporation of D amino-acids can significantly expand the peptide design space, which can in turn manifest in future biomaterial designs, especially antimicrobials.

Topological effects on the designability and bactericidal potency of antimicrobial peptides

New ideas and methods are being developed to generate highly designable small functional protein folds beyond the confines of natural structures, from secondary to quaternary level. Highly designable folds can have multiple sequence solutions, which are thermodynamically and kinetically stable. We have previously described how short syndiotactic helices can be exceptionally stable energetically, and how they can be used as a template for designing antibacterial agents. In this work, we have designed four syndiotactic, single turn, amphipathic; cationic 7-mer peptides which are the sequence and structural subset of earlier published 12-mer sequences. We examined the stability of the designed structures and its effects on the biological activity of such short peptide sequences. This was achieved by making objective comparisons between 12-mer and 7-mer sequences in terms of their antibacterial activity. Further, we investigated the mechanistic origins of clearly different bactericidal potency of single (7-mer) and double (12-mer) turn syndiotactic helices using molecular dynamics simulations. Our results suggest that conformationally constrained stable short double turn peptide scaffolds are highly designable, whereas single turn structures are more likely to be disordered. The stability of the designed peptide structure provides a platform for inclusion of multiple sequence variables and defined electrostatic fingerprints. Therefore, a stable peptide scaffold along with pre-defined electrostatic signatures can together be utilized for the design of novel antimicrobial peptides.

Electrostatic Variation of Haemagglutinin as a Hallmark of the Evolution of Avian Influenza Viruses

Avian influenza virus is a zoonotic agent that significantly impacts public health and the poultry industry. Monitoring viral evolution and spread is crucial for surveillance and tracing programmes, which are currently based on serological or DNA sequencing-phylogenetics analysis. However, virus-host interactions, antigenic drift and spreading of viral clades strongly depend on variation in the surface features of capsid proteins. We report here that in silico comparative structural analysis of haemagglutinin can reveal relevant evolutionary fingerprints, particularly when integrated with sequence-based analyses. Phylogenetic analyses of H9 viral strains from wild birds and poultry, performed with different methods, reliably led to clustering of viruses into five main groups. Subsequent comparison of structural features showed congruence between such clustering and surface electrostatic fingerprints. These latter fingerprints relate group-specific variations in electrostatic charges and isocontours to well-known haemagglutinin sites involved in the modulation of immune escape and host specificity. This work suggests that the integration of structural and sequence comparisons may enhance investigations of trends and relevant mechanisms in viral evolution.

Pandemic Avian Influenza and Intra/Interhaemagglutinin Subtype Electrostatic Variation among Viruses Isolated from Avian, Mammalian, and Human Hosts.

Host jump can result in deadly pandemic events when avian influenza A viruses broaden their host specificity and become able to infect mammals, including humans. Haemagglutinin—the major capsid protein in influenza A viruses—is subjected to high rate mutations, of which several occur at its “head”: the receptor-binding domain that mediates specific binding to host cell receptors. Such surface-changing mutations may lead to antigenically novel influenza A viruses hence in pandemics by host jump and in vaccine escape by antigenic drift. Changes in haemagglutinin surface electrostatics have been recently associated with antigenic drift and with clades evolution and spreading in H5N1 and H9N2 viruses. We performed a comparative analysis of haemagglutinin surface electrostatics to investigate clustering and eventual fingerprints among representative pandemic (H5 and H7) and nonpandemic (H4 and H6) avian influenza viral subtypes. We observed preferential sorting of viruses isolated from mammalian/human hosts among these electrostatic clusters of a subtype; however, sorting was not “100% specific” to the different clusters. Therefore, electrostatic fingerprints can help in understanding, but they cannot explain alone the host jumping mechanism.

Reaction Mechanism and Catalytic Fingerprint of Allantoin Racemase.

The stereospecific oxidative decomposition of urate into allantoin is the core of purine catabolism in many organisms. The spontaneous decomposition of upstream intermediates and the nonenzymatic racemization of allantoin lead to an accumulation of (R)-allantoin, because the enzymes converting allantoin into allantoate are specific for the (S) isomer. The enzyme allantoin racemase catalyzes the reversible conversion between the two allantoin enantiomers, thus ensuring the overall efficiency of the catabolic pathway and preventing allantoin accumulation. On the basis of recent crystallographic and biochemical evidence, allantoin racemase has been assigned to the family of cofactor-independent racemases, together with other amino acid racemases. A detailed computational investigation of allantoin racemase has been carried out to complement the available experimental data and to provide atomistic insight into the enzymatic action. Allantoin, the natural substrate of the enzyme, has been investigated at the quantum mechanical level, in order to rationalize its conformational and tautomeric equilibria, playing a key role in protein-ligand recognition and in the following catalytic steps. The reaction mechanism of the enzyme has been elucidated through quantum mechanics/molecular mechanics (QM/MM) calculations. The potential energy surface investigation, carried out at the QM/MM level, revealed a stepwise reaction mechanism. A pair of cysteine residues promotes the stereoinversion of a carbon atom of the ligand without the assistance of cofactors. Electrostatic fingerprint calculations are used to discuss the role of the active site residues in lowering the pKa of the substrate. The planar unprotonated intermediate is compared with the enolic allantoin tautomer observed in the active site of the crystallized enzyme. Finally, the enzymatic catalysis featured by allantoin racemase (AllR) is compared with that of other enzymes belonging to the same family.

Electrokinetic Analysis to Reveal Composition and Structure of Biohybrid Hydrogels

Biohybrid hydrogels combining electrically neutral synthetic polymers and highly anionic glycosaminoglycans (GAGs) offer exciting options for regenerative therapies as they allow for the electrostatic conjugation of various growth factors. Unraveling details of ionization and structure within such networks defines an important analytical challenge that requires the extension of current methodologies. Here, we present a mean-field approach to quantify the density of ionizable groups, GAG concentration, and cross-linking degree of such hydrogels based on experimental data from microslit electrokinetics and ellipsometry. An exemplary poly(ethylene glycol)-heparin system was analyzed to demonstrate how electrostatic fingerprints of hydrogels obtained by the introduced strategy can sensitively display composition and structure of the polymer networks.

The Effect of Electrostatic Fingerprint Visualization on Integrated Ballistic Identification Systems

Visualization of fingerprint corrosion on spent brass cartridge cases by the application of a high electrical potential and conducting carbon powder is becoming an accepted method of fingerprint enhancement. However, to date, no examination has been made of any effect this technique has on ballistic identification. To resolve this, images of the breech face and firing pin marks were captured on six plated nickel and six brass primer cup spent cartridge cases. Three nickel and three brass cases were then subjected to the application of a potential of +2500 V for a period of 1 min. The remaining cases were additionally subjected to the application of carbon powder. These latter cases were then washed to remove all traces of powder. Each case was recaptured with the same ballistic identification apparatus and imaging procedure. None of the twelve cases showed any visual difference after the application of the potential or conducting powder.

Imaging electrostatic fingerprints with implications for a forensic timeline

The electrical charge deposited by contact electrification due to a finger touching a thin insulating surface is imaged using an electric field microscopy system. It is based on an ultrahigh impedance electric potential sensor used as a non-contact raster scanning probe to measure surface charge density with a spatial resolution of up to 5 μm. Preliminary results are presented which yield two principle findings. First, they indicate that the spatial resolution of the fingerprint image is sufficient for identification purposes. Secondly, that the decay of the surface charge may be considered as a candidate method for the dating or sequencing of these electrical charge fingerprints. The decay of surface charge with time is well defined, largely material dependent, and may take many days. This intrinsic decay rate for the material may be quantified using the charge imaging system described in this paper and a known test charge. The measurement technique described is non-destructive, may be repeated without degradation of the sample, and does not preclude the subsequent use of other techniques such as DNA analysis or conventional latent fingerprint development.

Electrostatic properties in the catalytic site of papain: A possible regulatory mechanism for the reactivity of the ion pair.

We present an analysis of the electrostatic properties in the catalytic site of papain (EC 3.4.22.2), an archetype enzyme of the C1 cysteine proteinase family, and we investigate their possible role in the formation, stabilization and regulation of the Cys25((-))...His159((+)) catalytic ion pair. The electrostatic properties were computed using a reassociation method based in multicentered multipolar expansions obtained from ab initio quantum calculations of overlapping protein fragments. Solvent effects were introduced by coupling the use of multicentered multipolar expansions to two continuum boundary element methods to solve the Poisson and the linearized Poisson-Boltzmann equations. The electrostatic profile found in the proton transfer region of papain showed that this enzyme has a well-defined electrostatic environment to favor the formation and stabilization of the catalytic ion pair. The papain catalytic site electrostatic profile can be considered as an electrostatic fingerprint of the papain family with the following characteristics: (i) the presence of a net electric field highly aligned in the (Cys25)-SG-->(His159)-ND1 direction; (ii) the electrostatic profile has a saddle-point character; (iii) it is basically a local environmental effect. Furthermore, our analysis describes a possible regulatory mechanism (the E(SG-->ND1) attenuation effect) controlling the ion pair reactivity and permits to infer the Asp57 acidic residue as the most probable candidate to act as the electrostatic modulator.

The elusive atomic rationale for DNA base pair stability.

A systematic analysis of the electrostatic interaction between 27 natural DNA base pairs was carried out, based on ab initio correlated wave functions and the topology of the electron density. Using high rank multipole moments we show that the atomic partitioning of the interaction energy contains many substantial contributions between distant atoms. Profiles of cumulative energy versus internuclear distance show large fluctuations and provide an electrostatic fingerprint of the partitioning of interaction energy in a complex. A quantified comparison between each pair of energy profiles, one for each base pair, makes clear that there is no correlation between the total base pair interaction energy and the shape of the profile. In other words, base pairs with similar interaction energy are not stable for the same reasons in terms of atomic partitioning. In summary, simple rules to rationalize the pattern of energetic stability of naturally occurring base pairs in terms of subsets of atoms are elusive. Our work cautions against inappropriate use of Jorgensen's secondary interaction hypothesis.

A fast estimate of electrostatic group contributions to the free energy of protein-inhibitor binding.

Dissecting ligand-protein binding free energies in individual contributions of protein residues (which are referred to here as 'group contributions') is of significant importance. For example, such contributions could help in estimating the corresponding mutational effects and in studies of drug resistance problems. However, the meaning of group contributions is not always uniquely defined and the approximations for rapid estimates of such contributions are not well developed. In this paper, the nature of group contributions to binding free energy is examined, focusing particularly on electrostatic contributions which are expected to be well behaved. This analysis examines different definitions of group contributions; the 'relaxed' group contributions that represent the change in binding energy upon mutation of the given residue to glycine, and the 'non-relaxed' group contributions that represent the scaled Coulomb interaction between the given residue and the ligand. Both contributions are defined and evaluated by the linear response approximation (LRA) of the PDLD/ S method. The present analysis considers the binding of pepstatin to endothiapepsin and 23 of its mutants as a test case for a neutral ligand. The 'non-relaxed' group contributions of 15 endothiapepsin residues show significant peaks in the 'electrostatic fingerprint'. The residues that contribute to the electrostatic fingerprint are located in the binding site of endothiapepsin. They include the aspartic dyad (Asp32, Asp215) with adjacent residues and the flap region. Twelve of these 15 residues have a heavy atom distance of <3.75 A to pepstatin. The contributions of 8 (10) of these 12 residues can be reconciled with the calculated 'relaxed' group contributions where one allows the protein and solvent (solvent only) to relax upon mutation of the given residue to glycine. On the other hand, it was found that residues at the second 'solvation shell' can have relaxed contributions that are not captured by the non-relaxed approach. Hence, whereas residues with significant non-relaxed electrostatic contributions are likely to contribute to binding, residues with small non-relaxed contributions may still affect the binding energy. At any rate, it is established here that even in the case of uncharged inhibitors it is possible to use the non-relaxed electrostatic fingerprint to detect 'hot' residues that are responsible for binding. This is significant since some versions of the non-relaxed approximation are faster by several orders of magnitude than more rigorous approaches. The general applicability of this approach is outlined, emphasizing its potential in studies of drug resistance where it is crucial to have a rapid way of anticipating the effect of mutation on both drug binding and catalysis.

Possible Specificity of Cellular Interactions Due to Electrostatic Forces

It is known that the cells from a mixed population in a culture medium will finally segregate. This “social behavior” of the cells is a direct consequence of both physical and chemical interactions between cells. The physical forces involved in cell-cell interactions are considered to be the electrostatic, van der Waals, and very-short-range hydration forces.It has been believed until now that the electrostatic forces acting between identical cells were always repulsive and nonspecific. In the present work, we try to suggest that even the electrostatic forces could manifest a slightly specific character, so that a completely random Brownian motion of the cells could be influenced, corrected, and transformed into a “docking maneuver,” which could favor the specific interactions between cells suspended in a culture medium. The main point of our approach is that the peculiar patterns of electric charge distribution on the cell surfaces act like specific electrostatic fingerprints that could be responsible for the preferential interactions which precede cellular segregation (1).