The COVID-19 pandemic has changed the quality of life and economic systems all over the world, as the virus can be transmitted from human to human via air-droplets. Since the SARS-CoV-2 virus was first identified in 2019, the virus has naturally mutated over time. Southeast Asia is one of the areas in the world that has implemented various procedures and measures to slow down the disease outbreaks. The first cluster of COVID-19 was identified from the tourist-travel history, and then the diversity of coronavirus victims has posed a serious issue of human security on a massive scale. To evaluate whether or not naturally occurring mutations have strengthened the infectivity of SARS-CoV-2, we computed in silico the structural dynamics of the RBD-spike protein mutation enhancing ACE2-binding. When considering emerging variations in Southeast Asia, 14 dominant mutations were analyzed by applying the structural and energetic characterization using MD simulations. The ones in the RBD region displayed higher affinity to ACE2 due to the improved interfacial stability of the RBD β-strand surrounding the ACE2 across salt bridge hotspots. The binding hotspots and structurally conserved conformational-epitopes have been identified, which are deleterious for RBD mutation and ACE2 binding. We present an interactive visualization to facilitate the development of effective neutralizing agents for vaccination, prevention and treatment.

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Human bone morphogenetic protein-2 (hBMP-2) is a potent growth and differentiation factor for bone induction and regeneration. Recombinant hBMP-2 (rhBMP-2) was cloned and expressed as a soluble protein using E. coli-based expression system. A full-length gene encoding mature hBMP-2 was amplified by RT-PCR, cloned into an expression vector and expressed using SHuffle E. coli cells. The rhBMP-2 was successfully expressed as a soluble protein under the control of the lacUV5 and protein A promoters by IPTG induction. The rhBMP-2 fused with ZZ domain at its N-terminus was successively purified with a single step by using IgG Sepharose 6 fast flow affinity chromatography. Analysis of the purified protein on SDS-PAGE, Western blot analysis and LC-MS/MS, verified that the purified protein was rhBMP-2. The biological activity testing on hFOB 1.19 showed that rhBMP-2 had the ability to significantly induce cell proliferation in a dose dependent manner. ALP staining and activity assay also increased after rhBMP-2

Assembly and budding in the late-stage of human immunodeficiency virus type 1 (HIV-1) production relies on the polymerization of Gag protein at the inner leaflet of the plasma membrane. We previously generated an ankyrin repeat protein (Ank1D4) that specifically interacts with the CAp24 protein. This study aimed to improve the binding activity of Ank1D4 by generating two platforms for the Ank1D4 dimer. The design of these constructs featured a distinct orientation of monomeric Ank1D4 connected by a linker peptide (G S) . The binding surfaces in either dimer generated from the C-terminus of the Ank1D4 monomer linked with the N-terminus of another monomer (Ank1D4 ) or its inverted form (Ank1D4 ), similar to monomeric Ank1D4. The interaction of Ank1D4 with CAp24 from capture ELISA was significantly greater than that of Ank1D4 and the parental Ank1D4. The bifunctional characteristic of Ank1D4 was further demonstrated using sandwich ELISA. The binding kinetics of these ankyrins were evaluated using bio-layer interferometry analysis. The K of Ank1D4 , Ank1D4 and monomeric Ank1D4 was 3.5 nM, 53.7 nM, and 126.2 nM, respectively. The dynamics of the interdomain linker and the behavior of ankyrin dimers were investigated in silico. Upon the binding distance calculation from the candidate structures, the achievement in obtaining double active sites is more possible in Ank1D4 . The CD spectroscopic data indicated that secondary structure of dimer forms resemble Ank1D4 monomer α-helical content. This finding confers the strategy to generate dimer from rigid scaffold for acquiring the binding avidity.