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Sekhon, Simranjeet Singh,Ahn, Ji-Young,Shin, Woo-Ri,Ko, Jung Ho,Lee, Lyon,Dawes, Maisie,Tyler, John,Han, Janet,Kim, Sang Yong,Min, Jiho American Scientific Publishers 2017 Journal of nanoscience and nanotechnology Vol.17 No.12
<P>In this study, Pseudozyma jejuensis cutinase is immobilized on amino-functionalized magnetic supports by cross-linked enzyme aggregates (CLEA) for its use in bio-degradation of polycaprolactone (PCL). The amount of cutinase immobilized on the magnetic beads using glutaraldehyde as a coupling agent was measured to be 27 mg/g of beads with 82% recovery of activity after immobilization. Compared to the free enzyme, the immobilized cutinase showed the optimum pH at 1 unit higher (pH 8.0) and also retained its enzymatic activity at higher temperatures. There was 55% retention of cutinase activity after 50 consecutive reuses, indicating its stability and reusability in aqueous media. Moreover, the immobilized cutinase maintained more than 80% of its initial activity during 20 days storage period, while the free cutinase reduced to 48% of initial activity under same condition. In addition, the immobilized cutinase showed a more significant increase in biodegradability over the free cutinase when the immobilized cutinase was used to degrade PCL in a batch system. Higher thermal and storage stability, as well as good durability after repeated use of the immobilized cutinase CLEA, highlights its potential applicability as large scale continuous systems for the enzymatic degradation of PCL.</P>
Aptabody-aptatope interactions in aptablotting assays
Sekhon, S.,Um, H. J.,Shin, W. R.,Lee, S. H.,Min, J.,Ahn, J. Y.,Kim, Y. H. Royal Society of Chemistry 2017 Nanoscale Vol.9 No.22
<P>We demonstrate an aptablotting assay method that involves direct and indirect aptabody recognition. Nanoscale single-stranded DNA aptamers against GST and DIG-tags are utilized as aptabodies (GST-2 and DIG-1, respectively), and the GST-2 aptabody binding site, or aptatope, as predicted by a MOE-docking simulation of the protein-aptamer complex, shows the interaction of the GST-2 aptabody at the catalytically active region. The aptabody-aptatope interaction was evaluated by an in vitro enzyme inhibitory analysis. The binding capacity of the GST-2 aptabody was assessed by dot-blot, EMSA and SDS-PAGE/electroblot analyses, and the results showed that the aptabodies interact with both the native mono-/dimeric form and the denatured GST form on a membrane. The use of aptabodies can overcome the obstacles of current immunoblot assays, and these molecules are easily assessable via ELISA systems. Moreover, the hybridization of aptabodies and antibodies (hybrid-aptablotting) may have considerable impacts on the design of bioassay platforms.</P>
Porcine epidemic diarrhea (PED) infection, diagnosis and vaccination: A mini review
Sekhon, Simranjeet Singh,Nguyen, Phat-Loc,Ahn, Ji-Young,Lee, Kyeong-Ah,Lee, Lyon,Kim, Sang Yong,Yoon, Hobaek,Park, Jihoo,Ko, Jung Ho,Kim, Yang-Hoon Springer-Verlag 2016 Toxicology and Environmental Health Sciences Vol.8 No.5
Defining the copper binding aptamotif and aptamer integrated recovery platform (AIRP)
Sekhon, Simranjeet Singh,Lee, Sang-Hee,Lee, Kyeong-Ah,Min, Jiho,Lee, Byung-Tae,Kim, Kyoung-Woong,Ahn, Ji-Young,Kim, Yang-Hoon Royal Society of Chemistry 2017 Nanoscale Vol.9 No.8
<P>The potential copper binding sites in aptamers have been predicted on the basis of secondary structures and the binding affinity of aptamers with copper. Out of the 4 aptamers (Cu-A1 to Cu-A4) selected by SELEX and examined in the present study, the Cu-A2 aptamer shows the highest binding affinity to copper with the lowest KD value of 1.83 x 10(-11) M. In order to confirm the binding of copper to the proposed region, the binding affinity was experimentally validated using mutation and deletion analysis. We have confirmed that the high G-C pairing patterns and short stem-interval distance play important roles in copper binding. Aptamer specificity was also verified against diverse heavy metals. We also demonstrate an Aptamer Integrated Recovery Platform (AIRP) to recover copper from acidic mine drainage. AIRP can be easily regenerated at least 20 times without significant deterioration of the retrieval performance. To the best of our knowledge, AIRP is the first demonstration of copper specific recovery using aptamers. This can be scaled up and would have diverse applications in metal contaminated water treatment, recovery and as a potential biosensor for environmental analysis, monitoring, and risk assessment.</P>
Sekhon, Simranjeet Singh,Park, Jae-Min,Ahn, Ji-Young,Park, Tae Sun,Kwon, Soon-Dong,Kim, Yoon-Chul,Min, Jiho,Kim, Yang-Hoon 대한독성유전단백체학회 2014 Molecular & cellular toxicology Vol.10 No.2
The para-nitrobenzyl esterase (PNBE) was successfully immobilized onto electrospun polystyrene/poly(styrene-co-maleic anhydride) (PS/PSMA) nanofibers as cross-linked enzyme aggregates (CLEA). The maximum amount of PNBE immobilized on the PS-PSMA nanofibers was determined to be 56.4 mg/g of nanofibers. The variation of pH and temperature has been observed to affect the enzymatic activity of the free and immobilized PNBE-CLEA. The immobilized PNBE-CLEA has been observed to show improved stability as compared to the native one. PNBE-CLEA has been found to retain more than 63% activity after 60 consecutive reuses. Finally, the immobilized PNBE-CLEA presented a high decomposition efficiency of cephalexin tosylate PNB ester. The durability determination of PNBE-CLEA immobilized nanofibers showed 70% of de-esterification after 10 cycles. These results demonstrate that PNBE-CLEA immobilized nanofiber could be used as an economical and environment friendly system for the catalytic process of para-nitrobenzyl esters in pharmaceutical synthesis.
Structural and biochemical characterization of the cytosolic wheat cyclophilin TaCypA-1.
Sekhon, Simranjeet Singh,Kaur, Harsimran,Dutta, Tanima,Singh, Khushwant,Kumari, Sumita,Kang, Sunghyun,Park, Sung Goo,Park, Byoung Chul,Jeong, Dae Gwin,Pareek, Ashwani,Woo, Eui-Jeon,Singh, Prabhjeet,Yo Wiley-Blackwell 2013 Acta crystallographica. Section D, Biological crys Vol.69 No.4
<P>Cyclophilins belong to a family of proteins that bind to the immunosuppressive drug cyclosporin A (CsA). Several members of this protein family catalyze the cis-trans isomerization of peptide bonds preceding prolyl residues. The present study describes the biochemical and structural characteristics of a cytosolic cyclophilin (TaCypA-1) cloned from wheat (Triticum aestivum L.). Purified TaCypA-1 expressed in Escherichia coli showed peptidyl-prolyl cis-trans isomerase activity, which was inhibited by CsA with an inhibition constant of 78.3 nM. The specific activity and catalytic efficiency (kcat/Km) of the purified TaCypA-1 were 99.06 0.13 nmol s(-1) mg(-1) and 2.32 10(5) M(-1) s(-1), respectively. The structures of apo TaCypA-1 and the TaCypA-1-CsA complex were determined at 1.25 and 1.20?? resolution, respectively, using X-ray diffraction. Binding of CsA to the active site of TaCypA-1 did not result in any significant conformational change in the apo TaCypA-1 structure. This is consistent with the crystal structure of the human cyclophilin D-CsA complex reported at 0.96 ? resolution. The TaCypA-1 structure revealed the presence of a divergent loop of seven amino acids (48)KSGKPLH(54) which is a characteristic feature of plant cyclophilins. This study is the first to elucidate the structure of an enzymatically active plant cyclophilin which shows peptidyl-prolyl cis-trans isomerase activity and the presence of a divergent loop.</P>
Sekhon, Satpal Singh,Park, Jin-Soo,Choi, Young-Woo Royal Society of Chemistry 2010 Physical Chemistry Chemical Physics Vol.12 No.41
<P>Small-angle X-ray scattering (SAXS) technique has been used to study the evolution of ionomer peak in the recast Nafion membranes containing the ionic liquid 1-ethyl-3-methylimidazolium tetrafluoroborate (EMIBF<SUB>4</SUB>). To the best of our knowledge, this is the first report dealing with the study of membranes containing different concentrations of the ionic liquid (EMIBF<SUB>4</SUB>), from the dry (no ionic liquid) to the saturation (containing 70 wt% ionic liquid) state to understand the evolution of the ionomer peak and the formation of ionic aggregates in these anhydrous membranes. The small-angle scattering maximum (ionomer peak) has been observed to shift continuously toward lower scattering vector (<I>q</I>) values as the ionic liquid content increases. The ionic conductivity behavior for the membranes containing ionic liquid has been found to be closely related with the change of slope of the double logarithmic plot between the reciprocal of the position of the ionomer peak and the polymer weight fraction. The <I>q</I> region over which Porod's law has been obeyed in different membranes was initially narrow and has been observed to widen with an increase in the content of the ionic liquid.</P> <P>Graphic Abstract</P><P>Ionomer peak shifts to lower scattering vector values with an increase in ionic liquid content in anhydrous fuel cell membranes. <IMG SRC='http://pubs.rsc.org/services/images/RSCpubs.ePlatform.Service.FreeContent.ImageService.svc/ImageService/image/GA?id=c0cp00966k'> </P>