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Open Access Publications from the University of California

Scholarly Works (8 results)

  • Thesis
  • Peer Reviewed
UC Riverside Electronic Theses and Dissertations (2012)

Interactions between biomolecules are essential to biological function, and fundamentally understanding the forces that drives these interactions is of great medical importance. Molecular modeling approaches represent a powerful set of tools for gaining a mechanistic perspective of biomolecular interactions at the atomic level. Due to the complexities of biological environments, a diverse set of molecular modeling methods are often needed to capture various aspects of biomolecular function. Here we present a computational framework that utilizes established molecular modeling methods, such as molecular dynamics, Poisson-Boltzmann electrostatics, and small-molecule docking, as well as includes novel tools for elucidating the role of electrostatics in protein association. Our novel computational tool, Analysis of Electrostatics Similarity Of Proteins (AESOP), utilizes theoretical mutagenesis, electrostatic clustering, and electrostatic free energies of association to evaluate the role of each charged residue in protein association. The AESOP framework has been applied to various biomolecular systems, including barnase-bartstar, the gold-standard system for protein electrostatics, as well as for the successful design of novel SUMO-4 substrate analogs. One biological system that is of key interest is the complement immune system, which is an ancient component of innate immunity. The complement system is involved in the opsonization and clearance of foreign pathogens, and achieves its function through a cascade of protein-protein interactions largely driven by electrostatics. The key role of electrostatics in complement function is further evidenced by the existence of electrostatic "hot-spots" on the surface of complement proteins, and by the fact that pathogens utilize electrostatics in complement targeted evasion mechanisms. Over-activation of the complement system is implicated in numerous autoimmune and inflammatory diseases, and as a result anaphylatoxin receptor C5aR has become an important drug target. We have developed a conformationally-sampled pharmacophore model for known C5aR antagonists, which has utility in the design/evaluation of novel C5aR antagonists. We have also performed virtual screening based on a newly developed model of the interaction between C5aR and known potent antagonist PMX-53, and have identified a structurally diverse set of potential C5aR ligands. These studies define a computational framework that has utility in the analysis of many other protein targets.

    Cover page: Development and Applications of a Computational Framework for Protein and Drug Design
    • Article
    • Peer Reviewed
    UC Riverside Previously Published Works (2012)
    The interaction between complement fragment C3d and complement receptor 2 (CR2) is a key aspect of complement immune system activation, and is a component in a link between innate and adaptive immunities. The complement immune system is an ancient mechanism for defense, and can be found in species that have been on Earth for the last 600 million years. However, the link between the complement system and adaptive immunity, which is formed through the association of the B-cell co-receptor complex, including the C3d-CR2 interaction, is a much more recent adaptation. Human C3d and CR2 have net charges of -1 and +7 respectively, and are believed to have evolved favoring the role of electrostatics in their functions. To investigate the role of electrostatics in the function and evolution of human C3d and CR2, we have applied electrostatic similarity methods to identify regions of evolutionarily conserved electrostatic potential based on 24 homologues of complement C3d and 4 homologues of CR2. We also examine the effects of structural perturbation, as introduced through molecular dynamics and mutations, on spatial distributions of electrostatic potential to identify perturbation resistant regions, generated by so-called electrostatic "hot-spots". Distributions of electrostatic similarity based on families of perturbed structures illustrate the presence of electrostatic "hot-spots" at the two functional sites of C3d, while the surface of CR2 lacks electrostatic "hot-spots" despite its excessively positive nature. We propose that the electrostatic "hot-spots" of C3d have evolved to optimize its dual-functionality (covalently attaching to pathogen surfaces and interaction with CR2), which are both necessary for the formation B-cell co-receptor complexes. Comparison of the perturbation resistance of the electrostatic character of the homologues of C3d suggests that there was an emergence of a new role of electrostatics, and a transition in the function of C3d, after the divergence of jawless fish.
      Cover page: The Two Sides of Complement C3d: Evolution of Electrostatics in a Link between Innate and Adaptive Immunity
      • Article
      • Peer Reviewed
      UC Riverside Previously Published Works (2011)
      Electrostatic interactions are ubiquitous in proteins and dictate stability and function. In this review, we discuss several methods for the analysis of electrostatics in protein-protein interactions. We discuss alanine-scanning mutagenesis, Poisson-Boltzmann electrostatics, free energy calculations, electrostatic similarity distances, and hierarchical clustering of electrostatic potentials. Our recently developed computational framework, known as Analysis of Electrostatic Similarities Of Proteins (AESOP), incorporates these tools to efficiently elucidate the role of electrostatic potentials in protein interactions. We present the application of AESOP to several proteins and protein complexes, for which charge is purported to facilitate protein association. Specifically, we illustrate how recent work has shaped the formulation of electrostatic calculations, the correlation of electrostatic free energies and electrostatic potential clustering results with experimental binding and activity data, the pH dependence of protein stability and association, the design of mutant proteins with enhanced immunological activity, and how AESOP can expose deficiencies in structural models and experimental data. This integrative approach can be utilized to develop mechanistic models and to guide experimental studies by predicting mutations with desired physicochemical properties and function. Alteration of the electrostatic properties of proteins offers a basis for the design of proteins with optimized binding and activity.
        Cover page: Electrostatic Clustering and Free Energy Calculations Provide a Foundation for Protein Design and Optimization
        • Article
        • Peer Reviewed
        UC Riverside Previously Published Works (2012)
        The V3 loop of the glycoprotein 120 (gp120) is a contact point for cell entry of HIV-1 leading to infection. Despite sequence variability and lack of specific structure, the highly flexible V3 loop possesses a well-defined role in recognizing and selecting cell-bound coreceptors CCR5 and CXCR4 through a mechanism of charge complementarity. We have performed two independent molecular dynamics (MD) simulations to gain insights into the dynamic character of two V3 loops with slightly different sequences, but significantly different starting crystallographic structures. We have identified highly populated trajectory-specific salt bridges between oppositely charged stem residues Arg9 and Glu25 or Asp29. The two trajectories share nearly identical correlated motions within the simulations, despite their different overall structures. High occupancy salt bridges play a key role in the major cross-correlated motions in both trajectories, and may be responsible for transient structural stability in preparation for coreceptor binding. In addition, the two V3 loops visit conformations with similarities in spatial distributions of electrostatic potentials, despite their inherent flexibility, which may play a role in coreceptor recognition. It is plausible that cooperativity between overall electrostatic potential, charged residue interactions, and correlated motions could be associated with a coreceptor selection and binding.
          Cover page: Insights into the Structure, Correlated Motions, and Electrostatic Properties of Two HIV-1 gp120 V3 Loops
          • Article
          • Peer Reviewed
          UC Riverside Previously Published Works (2014)

          Background

          The complement protein C5a acts by primarily binding and activating the G-protein coupled C5a receptor C5aR (CD88), and is implicated in many inflammatory diseases. The cyclic hexapeptide PMX53 (sequence Ace-Phe-[Orn-Pro-dCha-Trp-Arg]) is a full C5aR antagonist of nanomolar potency, and is widely used to study C5aR function in disease.

          Results

          We construct for the first time molecular models for the C5aR:PMX53 complex without the a priori use of experimental constraints, via a computational framework of molecular dynamics (MD) simulations, docking, conformational clustering and free energy filtering. The models agree with experimental data, and are used to propose important intermolecular interactions contributing to binding, and to develop a hypothesis for the mechanism of PMX53 antagonism.

          Conclusion

          This work forms the basis for the design of improved C5aR antagonists, as well as for atomic-detail mechanistic studies of complement activation and function. Our computational framework can be widely used to develop GPCR-ligand structural models in membrane environments, peptidomimetics and other chemical compounds with potential clinical use.
            Cover page: Insights into the mechanism of C5aR inhibition by PMX53 via implicit solvent molecular dynamics simulations and docking
            • Article
            • Peer Reviewed
            UC Riverside Previously Published Works (2012)
            Abstract Background The V3 loop of the glycoprotein gp120 of HIV-1 plays an important role in viral entry into cells by utilizing as coreceptor CCR5 or CXCR4, and is implicated in the phenotypic tropisms of HIV viruses. It has been hypothesized that the interaction between the V3 loop and CCR5 or CXCR4 is mediated by electrostatics. We have performed hierarchical clustering analysis of the spatial distributions of electrostatic potentials and charges of V3 loop structures containing consensus sequences of HIV-1 subtypes. Results Although the majority of consensus sequences have a net charge of +3, the spatial distribution of their electrostatic potentials and charges may be a discriminating factor for binding and infectivity. This is demonstrated by the formation of several small subclusters, within major clusters, which indicates common origin but distinct spatial details of electrostatic properties. Some of this information may be present, in a coarse manner, in clustering of sequences, but the spatial details are largely lost. We show the effect of ionic strength on clustering of electrostatic potentials, information that is not present in clustering of charges or sequences. We also make correlations between clustering of electrostatic potentials and net charge, coreceptor selectivity, global prevalence, and geographic distribution. Finally, we interpret coreceptor selectivity based on the N6X7T8|S8X9 sequence glycosylation motif, the specific positive charge location according to the 11/24/25 rule, and the overall charge and electrostatic potential distribution. Conclusions We propose that in addition to the sequence and the net charge of the V3 loop of each subtype, the spatial distributions of electrostatic potentials and charges may also be important factors for receptor recognition and binding and subsequent viral entry into cells. This implies that the overall electrostatic potential is responsible for long-range recognition of the V3 loop with coreceptors CCR5/CXCR4, whereas the charge distribution contributes to the specific short-range interactions responsible for the formation of the bound complex. We also propose a scheme for coreceptor selectivity based on the sequence glycosylation motif, the 11/24/25 rule, and net charge.
              Cover page: Clustering of HIV-1 Subtypes Based on gp120 V3 Loop Electrostatic Properties
              • Article
              • Peer Reviewed
              UC Riverside Previously Published Works (2010)
              Lily stigma/style cysteine-rich adhesin (SCA), a plant lipid transfer protein (LTP) which is secreted into the extracellular matrix, functions in pollen tube guidance in fertilization. A gain-of-function mutant (ltp5-1) for Arabidopsis LTP5, an SCA-like molecule, was recently shown to display defects in sexual reproduction. In the current study, it is reported that ltp5-1 plants have dwarfed primary shoots, delayed hypocotyl elongation, various abnormal tissue fusions, and display multibranching. These mutant phenotypes in vegetative growth are recessive. No abnormality was found in ltp5-1/+ plants. In a phylogenetic analysis of plant LTPs, SCA-like Arabidopsis LTPs were classified with conventional plant LTPs. Homology modelling-based electrostatic similarity index (ESI) clustering was used to show diversity in spatial distributions of electrostatic potentials of SCA-like LTPs, suggestive of their various roles in interaction in the extracellular matrix space. β-Glucuronidase (GUS) analysis showed that SCA-like Arabidopsis LTP genes are diversely present in various tissues. LTP4 was found specifically in the guard cells and LTP6 in trichomes as well as in other tissues. LTP1 levels were specifically abundant in the stigma, and both LTP3 and LTP6 in the ovules. LTP2 and LTP4 gene levels were up-regulated in whole seedlings with 20% polyethylene glycol (PEG) and 300 mM NaCl treatments, respectively. LTP5 was up-regulated in the hypocotyl with 3 d dark growth conditions. LTP6 was specifically expressed in the tip of the cotyledon under drought stress conditions. The results suggest that SCA-like Arabidopsis LTPs are multifunctional, with diversified roles in plant growth and reproduction.
                Cover page: A multifaceted study of stigma/style cysteine-rich adhesin (SCA)-like Arabidopsis lipid transfer proteins (LTPs) suggests diversified roles for these LTPs in plant growth and reproduction
                • Article
                • Peer Reviewed
                LBL Publications (2018)
                Every two years groups worldwide participate in the Critical Assessment of Protein Structure Prediction (CASP) experiment to blindly test the strengths and weaknesses of their computational methods. CASP has significantly advanced the field but many hurdles still remain, which may require new ideas and collaborations. In 2012 a web-based effort called WeFold, was initiated to promote collaboration within the CASP community and attract researchers from other fields to contribute new ideas to CASP. Members of the WeFold coopetition (cooperation and competition) participated in CASP as individual teams, but also shared components of their methods to create hybrid pipelines and actively contributed to this effort. We assert that the scale and diversity of integrative prediction pipelines could not have been achieved by any individual lab or even by any collaboration among a few partners. The models contributed by the participating groups and generated by the pipelines are publicly available at the WeFold website providing a wealth of data that remains to be tapped. Here, we analyze the results of the 2014 and 2016 pipelines showing improvements according to the CASP assessment as well as areas that require further adjustments and research.
                  Cover page: An analysis and evaluation of the WeFold collaborative for protein structure prediction and its pipelines in CASP11 and CASP12
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