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RNA puzzles

We develop algorithms to predict the RNA structures of naturally occurring biomolecules and interactions with high accuracy. To carry out the RNA structure prediction calculations, we have been developing an automated and efficient program called 3dRNA (Scientific Reports, 2012; Nucleic Acids Research, 2017). We hypothesize that the organization of RNA structure is largely defined by topological constraints, and then build the RNA tertiary structure from the smallest secondary elements (SSEs) by using sequences and secondary structure information. The benchmark tests show that 3dRNA can give predictions with reasonable accuracy for RNAs of larger size and complex topology. We have continued to improve the 3dRNA automated structure prediction methodology by introducing a novel all-heavy-atom knowledge-based statistical potential, 3dRNAscore, to evaluate the predictions and to give better accuracy (Nucleic Acids Research, 2015). It turns out that 3dRNAscore performs better than current knowledge-based statistical potentials in identifying RNA native structures from a pool of structural decoys as well as ranking a tremendous amount of near-native RNA tertiary structures. The 3dRNA methodology is increasingly being used to solve structure-related problems in the RNA community.


Bimolecular mechanisms

Immune system The CD4+ and CD8+ T cell dichotomy is essential for effective cellular immunity. How individual T cell identity is established remains poorly understood. Here we show that the high-mobility group (HMG) transcription factors Tcf1 and Lef1 are essential for repressing CD4+ lineage–associated genes including Cd4, Foxp3 and Rorc in CD8+ T cells. Tcf1- and Lef1-deficient CD8+ T cells exhibit histone hyperacetylation, which can be ascribed to intrinsic histone deacetylase (HDAC) activity in Tcf1 and Lef1. Mutation of five conserved amino acids in the Tcf1 HDAC domain diminishes HDAC activity and the ability to suppress CD4+ lineage genes in CD8+ T cells. These findings reveal that sequence-specific transcription factors can utilize intrinsic HDAC activity to guard cell identity by repressing lineage-inappropriate genes. (Nature Immunology, 2016)


A new role for STAT3 as a regulator of chromatin topology The spatial organization of the genome is known to be important for regulation of gene transcription during normal development and in disease states. However, molecular mechanisms governing structural rearrangements in the genome are still poorly understood. We proposed a novel mechanism by which phosphorylated and unphosphorylated STAT3 and possibly other STAT proteins may regulate gene expression. The anticipated ability of P-STAT3 tetramers to bend DNA and U-STAT3 dimers to bring together distant DNA regions for DNA looping warrants further investigations as a potentially important, and thus far overlooked, mechanism by which STAT3 controls chromatin organization and gene expression. The proposed new function of STAT3 also emphasizes an important role of the NDs in STAT signaling. It suggests that inhibition of the ND may affect U-STAT dimerization and P-STAT tetramerization, thus preventing/inhibiting DNA looping/bending leading to changes in 3D chromatin structure and gene expression. Given the role STAT proteins play in regulation of immune responses, inflammation and cancer, the significance of further studies into architectural function of STAT proteins lies in discovering novel mechanisms by which STATs may regulate gene expression and designing novel approaches for manipulating them for therapeutic applications. (Transcription, 2013)


MD study of the mechanism of unfolding by MID1 Bbox1 domain mutations The zinc-binding Bbox1 domain in protein MID1, a member of the TRIM family of proteins, facilitates the ubiquitination of the catalytic subunit of protein phosphatase 2A and alpha4, a protein regulator of PP2A. The natural mutation of residue A130 to a valine or threonine disrupts substrate recognition and catalysis. While NMR data revealed the A130T mutant Bbox1 domain failed to coordinate both structurally essential zinc ions and resulted in an unfolded structure, the unfolding mechanism is unknown. MD simulations can be effective in understanding how the two mutations at position 130 perturb the protein structure. The accuracy in computational modeling and MD simulation has made it possible to study processes that are too fast to be observed experimentally. MD simulation analysis of the two A130T/V mutations revealed that residue A130 served as a hinge point and its mutation caused steric clashes that in turn increased the dielectric constant in one of the zinc-binding regions. The increased motion of residues on loop 1 disrupts the coordination of both zinc ions that are 13 Å apart. (PloS one, 2015)


Drug design

Break CDK2/Cyclin E1 Interface Allosterically with Small Peptides Most inhibitors of Cyclin-dependent kinase 2 (CDK2) target its ATP-binding pocket. It is difficult, however, to use this pocket to design very specific inhibitors because this catalytic pocket is highly conserved in the protein family of CDKs. In this project, we developed some short peptides targeting a noncatalytic pocket near the interface of the CDK2/Cyclin complex. Docking and molecular dynamics simulations were used to select the peptides, and detailed dynamical network analysis revealed that these peptides weaken the complex formation via allosteric interactions. Our Western Blot, Kinase assay and SPR assay experiments showed that upon binding to the noncatalytic pocket, these peptides break the CDK2/Cyclin complex partially and diminish its kinase activity in vitro. The binding affinity of these peptides measured by Surface Plasmon Resonance is low. (PloS one, 2014)


Design of Common Bean Lectin Inhibitor and Its Hemagglutination Activity Common bean (Phaseolus vulgaris L.), rich in protein and low in fat, is one of the most important legume crops in the world. However, consumption of improperly prepared common bean can lead to food poisoning. Previous studies have largely attributed this toxic effect to the high content in lectin, a plant protein that binds specifically to carbohydrate or carbohydrate structures displayed on cell surface. Since most lectin forms a dimer complex for biological functions, thus a short peptide was designed to break the dimer interface. Detailed dynamical network and structural characteristic analysis were performed to select this peptide. In vitro hemagglutination assay showed that this peptide, upon binding to lectin, disrupts the dimer formation partially and weakens the hemagglutination effect. Taken together, a novel peptide inhibitor was designed whose potency and specificity can be further optimized for anti-hemagglutination and food safety applications. (Chem J Chinese U, 2017)


Machine Learning or Deep Learning

Network Analysis Reveals the Recognition Mechanism for Dimer Formation of Bulb-type Lectins The bulb-type lectins are proteins consist of three sequential beta-sheet subdomains that bind to speci c carbohydrates to perform certain biological functions. The active states of most bulb-type lectins are dimeric and it is thus important to elucidate the short- and long-range recognition mechanism for this dimer formation. To do so, we perform comparative sequence analysis for the single- and double-domain bulb-type lectins abundant in plant genomes. In contrast to the dimer complex of two single-domain lectins formed via protein-protein interactions, the double-domain lectin fuses two single-domain proteins into one protein with a short linker and requires only short-range interactions because its two single domains are always in close proximity. Sequence analysis demonstrates that the highly variable but coevolving polar residues at the interface of dimeric bulb-type lectins are largely absent in the double-domain bulb-type lectins. Moreover, network analysis on bulb-type lectin proteins show that these same polar residues have high closeness scores and thus serve as hubs with strong connections to all other residues. Taken together, we propose a potential mechanism for this lectin complex formation where coevolving polar residues of high closeness are responsible for long-range recognition. (Scientific Reports, 2017)

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