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1.  Structural and Biophysical Characterization of Bacillus thuringiensis Insecticidal Proteins Cry34Ab1 and Cry35Ab1 
PLoS ONE  2014;9(11):e112555.
Bacillus thuringiensis strains are well known for the production of insecticidal proteins upon sporulation and these proteins are deposited in parasporal crystalline inclusions. The majority of these insect-specific toxins exhibit three domains in the mature toxin sequence. However, other Cry toxins are structurally and evolutionarily unrelated to this three-domain family and little is known of their three dimensional structures, limiting our understanding of their mechanisms of action and our ability to engineer the proteins to enhance their function. Among the non-three domain Cry toxins, the Cry34Ab1 and Cry35Ab1 proteins from B. thuringiensis strain PS149B1 are required to act together to produce toxicity to the western corn rootworm (WCR) Diabrotica virgifera virgifera Le Conte via a pore forming mechanism of action. Cry34Ab1 is a protein of ∼14 kDa with features of the aegerolysin family (Pfam06355) of proteins that have known membrane disrupting activity, while Cry35Ab1 is a ∼44 kDa member of the toxin_10 family (Pfam05431) that includes other insecticidal proteins such as the binary toxin BinA/BinB. The Cry34Ab1/Cry35Ab1 proteins represent an important seed trait technology having been developed as insect resistance traits in commercialized corn hybrids for control of WCR. The structures of Cry34Ab1 and Cry35Ab1 have been elucidated to 2.15 Å and 1.80 Å resolution, respectively. The solution structures of the toxins were further studied by small angle X-ray scattering and native electrospray ion mobility mass spectrometry. We present here the first published structure from the aegerolysin protein domain family and the structural comparisons of Cry34Ab1 and Cry35Ab1 with other pore forming toxins.
PMCID: PMC4229197  PMID: 25390338
Molecular pharmaceutics  2013;10(10):10.1021/mp3007242.
5P12-RANTES is a recently developed chemokine analog that has shown high level protection from SHIV infection in macaques. However, the feasibility of using 5P12-RANTES as a long term HIV prevention agent has not been explored partially due to the lack of available delivery devices that can easily be modified for long-term release profiles. Glycosaminoglycans (GAGs) have been known for their affinity for various cytokines and chemokines, including native RANTES, or CCL5. In this work, we investigated used of GAGs in generating a chemokine drug delivery device. Initial studies used surface plasmon resonance analysis to characterize and compare the affinities of different GAGs to 5P12-RANTES. These different GAGs were then incorporated into drug delivery polymeric hydrogels to engineer sustained release of the chemokines. In vitro release studies of 5P12-RANTES from the resulting polymers were performed and we found that 5P12-RANTES release from these polymers can be controlled by the amount and type of GAG incorporated. Polymer disks containing GAGs with stronger affinity to 5P12-RANTES resulted in more sustained, and longer term release than did polymer disks containing GAGs with weaker 5P12-RANTES affinity. Similar trends were observed by varying the amount of GAGs incorporated into the delivery system. 5P12-RANTES released from these polymers demonstrated good levels of CCR5 blocking, retaining activity even after 30 days of incubation.
PMCID: PMC3886841  PMID: 23859720
HIV; drug delivery; prevention; microbicide; chemokine; CCL5; CCR5; glycosaminoglycans; heparin
3.  Therapeutic use of PDZ protein-protein interaction antagonism 
Drug news & perspectives  2008;21(3):137-141.
PDZ domains are important interaction modules in many intracellular pathways and abnormal activations of many of those pathways lead to diseases, including several types of cancer. The domains are characterized by the ability to recognize the extreme COOH-terminus of target proteins, such as G protein-coupled receptors and ion channels. Because PDZ protein-protein interaction is a key factor in the function of cellular pathways and signal transmission in those pathways, developing small-molecule inhibitors to compete with PDZ targets is very attractive in dissecting molecular mechanisms and formulating pharmaceutical agents. Moreover, there is a growing interest in developing small-molecule drugs to block signaling within cells. The modulation of PDZ-involved interactions in cells might be an approach to target the G protein-coupled receptors and ion channels, which are among the most important classes of drug targets in the pharmaceutical industry today. Here, we review recent progress in the development of small-molecule PDZ inhibitors, and especially focus on two PDZ domain-containing target proteins, postsynaptic density 95 and dishevelled.
PMCID: PMC4055467  PMID: 18560611
4.  Identification Of Small Molecule TRABID Deubiquitinase Inhibitors By Computation-Based Virtual Screen 
BMC Chemical Biology  2012;12:4.
Wnt/β-catenin-mediated gene transcription plays important roles in a wide range of biological and pathophysiological processes including tumorigenesis where β-catenin-mediated transcription activity frequently elevates. TRABID, a deubiquitinase, was shown to have a positive Wnt/β-catenin-mediated gene transcription and hence holds a promise as a putative anti-cancer target.
In this study, we used a combination of structure based virtual screening and an in vitro deubiquitinase (DUB) assay to identify several small molecules that inhibit TRABID DUB activity. However, these inhibitors failed to show inhibitory effects on β-catenin-mediated gene transcription. In addition, expression of TRABID shRNAs, wildtype TRABID, or the DUB activity-deficient mutant showed little effects on β-catenin-mediated gene transcription.
TRABID may not be a critical component in canonical Wnt/β-catenin signal transduction or that a minute amount of this protein is sufficient for its role in regulating Wnt activity.
PMCID: PMC3475094  PMID: 22584113
6.  Identification of Tripeptides Recognized by the PDZ domain of Dishevelled 
Bioorganic & medicinal chemistry  2009;17(4):1701-1708.
The development of inhibitors of Dishevelled (Dvl) PDZ protein-protein interactions attracts attention due to a possible application in drug discovery and development. Using nuclear magnetic resonance (NMR) spectroscopy, we found that a tripeptide VVV binds to the PDZ domain of Dvl, which is a key component involved in Wnt signaling. Using a computational approach calculating the binding free energy of the complexes of the Dvl PDZ domain and each of the tripeptides VXV (X: any amino acid residue except Pro), we found that a tripeptide VWV had the highest binding affinity. Consistent with the computational result, experimental results showed that the binding of the tripeptide VWV to the Dvl PDZ domain was stronger than that of the tripeptide VVV. The binding affinity of the tripeptide VWV was comparable to that of the organic molecule NSC668036, which was the first identified Dvl PDZ inhibitor. The three-dimensional structure of the complex Dvl1 PDZ/VWV was determined to investigate the role of the energetically favorable W(−1) residue in binding. These interactions were also explored by using molecular dynamic simulation and the molecular mechanics Poisson-Boltzmann surface area method. Taken together, these two tripeptides may be used as modulators of Wnt signaling or as a scaffold to optimize an antagonist for targeting Dvl1 PDZ protein-protein interaction.
PMCID: PMC2713185  PMID: 19157887
binding free energy; protein-ligand interaction; ligand computational screening; NMR; fluorescence spectroscopy

Results 1-6 (6)