Publications by authors named "Doyoun Kim"

38 Publications

SLC6A20 transporter: a novel regulator of brain glycine homeostasis and NMDAR function.

EMBO Mol Med 2021 Feb 11;13(2):e12632. Epub 2021 Jan 11.

Center for Synaptic Brain Dysfunctions, Institute for Basic Science (IBS), Daejeon, Korea.

Glycine transporters (GlyT1 and GlyT2) that regulate levels of brain glycine, an inhibitory neurotransmitter with co-agonist activity for NMDA receptors (NMDARs), have been considered to be important targets for the treatment of brain disorders with suppressed NMDAR function such as schizophrenia. However, it remains unclear whether other amino acid transporters expressed in the brain can also regulate brain glycine levels and NMDAR function. Here, we report that SLC6A20A, an amino acid transporter known to transport proline based on in vitro data but is understudied in the brain, regulates proline and glycine levels and NMDAR function in the mouse brain. SLC6A20A transcript and protein levels were abnormally increased in mice carrying a mutant PTEN protein lacking the C terminus through enhanced β-catenin binding to the Slc6a20a gene. These mice displayed reduced extracellular levels of brain proline and glycine and decreased NMDAR currents. Elevating glycine levels back to normal ranges by antisense oligonucleotide-induced SLC6A20 knockdown, or the competitive GlyT1 antagonist sarcosine, normalized NMDAR currents and repetitive climbing behavior observed in these mice. Conversely, mice lacking SLC6A20A displayed increased extracellular glycine levels and NMDAR currents. Lastly, both mouse and human SLC6A20 proteins mediated proline and glycine transports, and SLC6A20 proteins could be detected in human neurons. These results suggest that SLC6A20 regulates proline and glycine homeostasis in the brain and that SLC6A20 inhibition has therapeutic potential for brain disorders involving NMDAR hypofunction.
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http://dx.doi.org/10.15252/emmm.202012632DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7863395PMC
February 2021

Clmp Regulates AMPA and Kainate Receptor Responses in the Neonatal Hippocampal CA3 and Kainate Seizure Susceptibility in Mice.

Front Synaptic Neurosci 2020 21;12:567075. Epub 2020 Dec 21.

Center for Synaptic Brain Dysfunctions, Institute for Basic Science, Daejeon, South Korea.

Synaptic adhesion molecules regulate synapse development through trans-synaptic adhesion and assembly of diverse synaptic proteins. Many synaptic adhesion molecules positively regulate synapse development; some, however, exert negative regulation, although such cases are relatively rare. In addition, synaptic adhesion molecules regulate the amplitude of post-synaptic receptor responses, but whether adhesion molecules can regulate the kinetic properties of post-synaptic receptors remains unclear. Here we report that Clmp, a homophilic adhesion molecule of the Ig domain superfamily that is abundantly expressed in the brain, reaches peak expression at a neonatal stage (week 1) and associates with subunits of AMPA receptors (AMPARs) and kainate receptors (KARs). deletion in mice increased the frequency and amplitude of AMPAR-mediated miniature excitatory post-synaptic currents (mEPSCs) and the frequency, amplitude, and decay time constant of KAR-mediated mEPSCs in hippocampal CA3 neurons. deletion had minimal impacts on evoked excitatory synaptic currents at mossy fiber-CA3 synapses but increased extrasynaptic KAR, but not AMPAR, currents, suggesting that Clmp distinctly inhibits AMPAR and KAR responses. Behaviorally, deletion enhanced novel object recognition and susceptibility to kainate-induced seizures, without affecting contextual or auditory cued fear conditioning or pattern completion-based contextual fear conditioning. These results suggest that Clmp negatively regulates hippocampal excitatory synapse development and AMPAR and KAR responses in the neonatal hippocampal CA3 as well as object recognition and kainate seizure susceptibility in mice.
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http://dx.doi.org/10.3389/fnsyn.2020.567075DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7779639PMC
December 2020

Therapeutic Strategies Against COVID-19 and Structural Characterization of SARS-CoV-2: A Review.

Front Microbiol 2020 14;11:1723. Epub 2020 Jul 14.

Center for Convergence for Emerging Virus Infection, Korea Research Institute of Chemical Technology (KRICT), Daejeon, South Korea.

The novel coronavirus, SARS-CoV-2, or 2019-nCoV, which originated in Wuhan, Hubei province, China in December 2019, is a grave threat to public health worldwide. A total of 3,672,238 confirmed cases of coronavirus disease 2019 (COVID-19) and 254,045 deaths were reported globally up to May 7, 2020. However, approved antiviral agents for the treatment of patients with COVID-19 remain unavailable. Drug repurposing of approved antivirals against other viruses such as HIV or Ebola virus is one of the most practical strategies to develop effective antiviral agents against SARS-CoV-2. A combination of repurposed drugs can improve the efficacy of treatment, and structure-based drug design can be employed to specifically target SARS-CoV-2. This review discusses therapeutic strategies using promising antiviral agents against SARS-CoV-2. In addition, structural characterization of potentially therapeutic viral or host cellular targets associated with COVID-19 have been discussed to refine structure-based drug design strategies.
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http://dx.doi.org/10.3389/fmicb.2020.01723DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7381222PMC
July 2020

A novel de novo heterozygous DYRK1A mutation causes complete loss of DYRK1A function and developmental delay.

Sci Rep 2020 06 17;10(1):9849. Epub 2020 Jun 17.

Division of Medical Genetics and Metabolism, Department of Pediatrics, Pusan National University Children's Hospital, Pusan National University School of Medicine, 20 Geumo-ro, Mulgeum-eup, Yangsan-si, Gyeongnam, 50612, Republic of Korea.

Dual-specificity tyrosine phosphorylation-regulated kinase 1 A (DYRK1A) is essential for human development, and DYRK1A haploinsufficiency is associated with a recognizable developmental syndrome and variable clinical features. Here, we present a patient with DYRK1A haploinsufficiency syndrome, including facial dysmorphism, delayed motor development, cardiovascular system defects, and brain atrophy. Exome sequencing identified a novel de novo heterozygous mutation of the human DYRK1A gene (c.1185dup), which generated a translational termination codon and resulted in a C-terminally truncated protein (DYRK1A-E396ter). To study the molecular effect of this truncation, we generated mammalian cell and Drosophila models that recapitulated the DYRK1A protein truncation. Analysis of the structure and deformation energy of the mutant protein predicted a reduction in protein stability. Experimentally, the mutant protein was efficiently degraded by the ubiquitin-dependent proteasome pathway and was barely detectable in mammalian cells. More importantly, the mutant kinase was intrinsically inactive and had little negative impact on the wild-type protein. Similarly, the mutant protein had a minimal effect on Drosophila phenotypes, confirming its loss-of-function in vivo. Together, our results suggest that the novel heterozygous mutation of DYRK1A resulted in loss-of-function of the kinase activity of DYRK1A and may contribute to the developmental delay observed in the patient.
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http://dx.doi.org/10.1038/s41598-020-66750-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7299959PMC
June 2020

Early correction of synaptic long-term depression improves abnormal anxiety-like behavior in adult GluN2B-C456Y-mutant mice.

PLoS Biol 2020 04 30;18(4):e3000717. Epub 2020 Apr 30.

Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Korea.

Extensive evidence links Glutamate receptor, ionotropic, NMDA2B (GRIN2B), encoding the GluN2B/NR2B subunit of N-methyl-D-aspartate receptors (NMDARs), with various neurodevelopmental disorders, including autism spectrum disorders (ASDs), but the underlying mechanisms remain unclear. In addition, it remains unknown whether mutations in GluN2B, which starts to be expressed early in development, induces early pathophysiology that can be corrected by early treatments for long-lasting effects. We generated and characterized Grin2b-mutant mice that carry a heterozygous, ASD-risk C456Y mutation (Grin2b+/C456Y). In Grin2b+/C456Y mice, GluN2B protein levels were strongly reduced in association with decreased hippocampal NMDAR currents and NMDAR-dependent long-term depression (LTD) but unaltered long-term potentiation, indicative of mutation-induced protein degradation and LTD sensitivity. Behaviorally, Grin2b+/C456Y mice showed normal social interaction but exhibited abnormal anxiolytic-like behavior. Importantly, early, but not late, treatment of young Grin2b+/C456Y mice with the NMDAR agonist D-cycloserine rescued NMDAR currents and LTD in juvenile mice and improved anxiolytic-like behavior in adult mice. Therefore, GluN2B-C456Y haploinsufficiency decreases GluN2B protein levels, NMDAR-dependent LTD, and anxiety-like behavior, and early activation of NMDAR function has long-lasting effects on adult mouse behavior.
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http://dx.doi.org/10.1371/journal.pbio.3000717DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7217483PMC
April 2020

Presynaptic PTPσ regulates postsynaptic NMDA receptor function through direct adhesion-independent mechanisms.

Elife 2020 03 6;9. Epub 2020 Mar 6.

Department of Biological Sciences, KAIST, Daejeon, Republic of Korea.

Synaptic adhesion molecules regulate synapse development and function. However, whether and how presynaptic adhesion molecules regulate postsynaptic NMDAR function remains largely unclear. Presynaptic LAR family receptor tyrosine phosphatases (LAR-RPTPs) regulate synapse development through mechanisms that include trans-synaptic adhesion; however, whether they regulate postsynaptic receptor functions remains unknown. Here we report that presynaptic PTPσ, a LAR-RPTP, enhances postsynaptic NMDA receptor (NMDAR) currents and NMDAR-dependent synaptic plasticity in the hippocampus. This regulation does not involve trans-synaptic adhesions of PTPσ, suggesting that the cytoplasmic domains of PTPσ, known to have tyrosine phosphatase activity and mediate protein-protein interactions, are important. In line with this, phosphotyrosine levels of presynaptic proteins, including neurexin-1, are strongly increased in PTPσ-mutant mice. Behaviorally, PTPσ-dependent NMDAR regulation is important for social and reward-related novelty recognition. These results suggest that presynaptic PTPσ regulates postsynaptic NMDAR function through trans-synaptic and direct adhesion-independent mechanisms and novelty recognition in social and reward contexts.
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http://dx.doi.org/10.7554/eLife.54224DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7069723PMC
March 2020

A TBR1-K228E Mutation Induces Upregulation, Altered Cortical Distribution of Interneurons, Increased Inhibitory Synaptic Transmission, and Autistic-Like Behavioral Deficits in Mice.

Front Mol Neurosci 2019 9;12:241. Epub 2019 Oct 9.

College of Pharmacy, Yeongnam University, Gyeongsan, South Korea.

Mutations in , a high-confidence ASD (autism spectrum disorder)-risk gene encoding the transcriptional regulator TBR1, have been shown to induce diverse ASD-related molecular, synaptic, neuronal, and behavioral dysfunctions in mice. However, whether mutations derived from autistic individuals cause similar dysfunctions in mice remains unclear. Here we generated and characterized mice carrying the TBR1-K228E mutation identified in human ASD and identified various ASD-related phenotypes. In heterozygous mice carrying this mutation ( mice), levels of the TBR1-K228E protein, which is unable to bind target DNA, were strongly increased. RNA-Seq analysis of the embryonic brain indicated significant changes in the expression of genes associated with neurons, astrocytes, ribosomes, neuronal synapses, and ASD risk. The neocortex also displayed an abnormal distribution of parvalbumin-positive interneurons, with a lower density in superficial layers but a higher density in deep layers. These changes were associated with an increase in inhibitory synaptic transmission in layer 6 pyramidal neurons that was resistant to compensation by network activity. Behaviorally, mice showed decreased social interaction, increased self-grooming, and modestly increased anxiety-like behaviors. These results suggest that the human heterozygous TBR1-K228E mutation induces ASD-related transcriptomic, protein, neuronal, synaptic, and behavioral dysfunctions in mice.
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http://dx.doi.org/10.3389/fnmol.2019.00241DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6797848PMC
October 2019

Reduced CYFIP2 Stability by Arg87 Variants Causing Human Neurological Disorders.

Ann Neurol 2019 11 4;86(5):803-805. Epub 2019 Oct 4.

Department of Neuroscience, College of Medicine, Korea University, Seoul.

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http://dx.doi.org/10.1002/ana.25598DOI Listing
November 2019

Mice Carrying the Human Q321R Mutation Display Enhanced Self-Grooming, Abnormal Electroencephalogram Patterns, and Suppressed Neuronal Excitability and Seizure Susceptibility.

Front Mol Neurosci 2019 18;12:155. Epub 2019 Jun 18.

Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon, South Korea.

Shank3, a postsynaptic scaffolding protein involved in regulating excitatory synapse assembly and function, has been implicated in several brain disorders, including autism spectrum disorders (ASD), Phelan-McDermid syndrome, schizophrenia, intellectual disability, and mania. Here we generated and characterized a knock-in mouse line carrying the Q321R mutation ( mice) identified in a human individual with ASD that affects the ankyrin repeat region (ARR) domain of the Shank3 protein. Homozygous mice show a selective decrease in the level of Shank3a, an ARR-containing protein variant, but not other variants. CA1 pyramidal neurons in the hippocampus show decreased neuronal excitability but normal excitatory and inhibitory synaptic transmission. Behaviorally, mice show moderately enhanced self-grooming and anxiolytic-like behavior, but normal locomotion, social interaction, and object recognition and contextual fear memory. In addition, these mice show abnormal electroencephalogram (EEG) patterns and decreased susceptibility to induced seizures. These results indicate that the Q321R mutation alters Shank3 protein stability, neuronal excitability, repetitive and anxiety-like behavior, EEG patterns, and seizure susceptibility in mice.
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http://dx.doi.org/10.3389/fnmol.2019.00155DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6591539PMC
June 2019

Haploinsufficiency in Mice Suppresses Hippocampal Neuronal Excitability, Excitatory Synaptic Drive, and Long-Term Potentiation, and Spatial Learning and Memory.

Front Mol Neurosci 2019 4;12:145. Epub 2019 Jun 4.

Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea.

Nav1.2, a voltage-gated sodium channel subunit encoded by the gene, has been implicated in various brain disorders, including epilepsy, autism spectrum disorder, intellectual disability, and schizophrenia. Nav1.2 is known to regulate the generation of action potentials in the axon initial segment and their propagation along axonal pathways. Nav1.2 also regulates synaptic integration and plasticity by promoting back-propagation of action potentials to dendrites, but whether Nav1.2 deletion in mice affects neuronal excitability, synaptic transmission, synaptic plasticity, and/or disease-related animal behaviors remains largely unclear. Here, we report that mice heterozygous for the gene ( mice) show decreased neuronal excitability and suppressed excitatory synaptic transmission in the presence of network activity in the hippocampus. In addition, mice show suppressed hippocampal long-term potentiation (LTP) in association with impaired spatial learning and memory, but show largely normal locomotor activity, anxiety-like behavior, social interaction, repetitive behavior, and whole-brain excitation. These results suggest that Nav1.2 regulates hippocampal neuronal excitability, excitatory synaptic drive, LTP, and spatial learning and memory in mice.
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http://dx.doi.org/10.3389/fnmol.2019.00145DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6582764PMC
June 2019

Early Correction of N-Methyl-D-Aspartate Receptor Function Improves Autistic-like Social Behaviors in Adult Shank2 Mice.

Biol Psychiatry 2019 04 9;85(7):534-543. Epub 2018 Oct 9.

Department of Biological Sciences, South Korea; Center for Synaptic Brain Dysfunctions, Institute for Basic Science, South Korea. Electronic address:

Background: Autism spectrum disorder involves neurodevelopmental dysregulations that lead to visible symptoms at early stages of life. Many autism spectrum disorder-related mechanisms suggested by animal studies are supported by demonstrated improvement in autistic-like phenotypes in adult animals following experimental reversal of dysregulated mechanisms. However, whether such mechanisms also act at earlier stages to cause autistic-like phenotypes is unclear.

Methods: We used Shank2 mice carrying a mutation identified in human autism spectrum disorder (exons 6 and 7 deletion) and combined electrophysiological and behavioral analyses to see whether early pathophysiology at pup stages is different from late pathophysiology at juvenile and adult stages and whether correcting early pathophysiology can normalize late pathophysiology and abnormal behaviors in juvenile and adult mice.

Results: Early correction of a dysregulated mechanism in young mice prevents manifestation of autistic-like social behaviors in adult mice. Shank2 mice, known to display N-methyl-D-aspartate receptor (NMDAR) hypofunction and autistic-like behaviors at postweaning stages after postnatal day 21 (P21), show the opposite synaptic phenotype-NMDAR hyperfunction-at an earlier preweaning stage (∼P14). Moreover, this NMDAR hyperfunction at P14 rapidly shifts to NMDAR hypofunction after weaning (∼P24). Chronic suppression of the early NMDAR hyperfunction by the NMDAR antagonist memantine (P7-P21) prevents NMDAR hypofunction and autistic-like social behaviors from manifesting at later stages (∼P28 and P56).

Conclusions: Early NMDAR hyperfunction leads to late NMDAR hypofunction and autistic-like social behaviors in Shank2 mice, and early correction of NMDAR dysfunction has the long-lasting effect of preventing autistic-like social behaviors from developing at later stages.
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http://dx.doi.org/10.1016/j.biopsych.2018.09.025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6420362PMC
April 2019

Sequence preference and structural heterogeneity of BZ junctions.

Nucleic Acids Res 2018 11;46(19):10504-10513

Department of Molecular Cell Biology, Institute for Antimicrobial Research and Therapeutics, Sungkyunkwan University School of Medicine, Suwon 16419, Korea.

BZ junctions, which connect B-DNA to Z-DNA, are necessary for local transformation of B-DNA to Z-DNA in the genome. However, the limited information on the junction-forming sequences and junction structures has led to a lack of understanding of the structural diversity and sequence preferences of BZ junctions. We determined three crystal structures of BZ junctions with diverse sequences followed by spectroscopic validation of DNA conformation. The structural features of the BZ junctions were well conserved regardless of sequences via the continuous base stacking through B-to-Z DNA with A-T base extrusion. However, the sequence-dependent structural heterogeneity of the junctions was also observed in base step parameters that are correlated with steric constraints imposed during Z-DNA formation. Further, circular dichroism and fluorescence-based analysis of BZ junctions revealed that a base extrusion was only found at the A-T base pair present next to a stable dinucleotide Z-DNA unit. Our findings suggest that Z-DNA formation in the genome is influenced by the sequence preference for BZ junctions.
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http://dx.doi.org/10.1093/nar/gky784DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6212838PMC
November 2018

Lrfn2-Mutant Mice Display Suppressed Synaptic Plasticity and Inhibitory Synapse Development and Abnormal Social Communication and Startle Response.

J Neurosci 2018 06 24;38(26):5872-5887. Epub 2018 May 24.

Center for Synaptic Brain Dysfunctions, Institute for Basic Science (IBS), Daejeon 34141, Korea,

SALM1 (SALM (synaptic adhesion-like molecule), also known as LRFN2 (leucine rich repeat and fibronectin type III domain containing), is a postsynaptic density (PSD)-95-interacting synaptic adhesion molecule implicated in the regulation of NMDA receptor (NMDAR) clustering largely based on data, although its functions remain unclear. Here, we found that mice lacking SALM1/LRFN2 ( mice) show a normal density of excitatory synapses but altered excitatory synaptic function, including enhanced NMDAR-dependent synaptic transmission but suppressed NMDAR-dependent synaptic plasticity in the hippocampal CA1 region. Unexpectedly, SALM1 expression was detected in both glutamatergic and GABAergic neurons and CA1 pyramidal neurons showed decreases in the density of inhibitory synapses and the frequency of spontaneous inhibitory synaptic transmission. Behaviorally, ultrasonic vocalization was suppressed in pups separated from their mothers and acoustic startle was enhanced, but locomotion, anxiety-like behavior, social interaction, repetitive behaviors, and learning and memory were largely normal in adult male mice. These results suggest that SALM1/LRFN2 regulates excitatory synapse function, inhibitory synapse development, and social communication and startle behaviors in mice. Synaptic adhesion molecules regulate synapse development and function, which govern neural circuit and brain functions. The SALM/LRFN (synaptic adhesion-like molecule/leucine rich repeat and fibronectin type III domain containing) family of synaptic adhesion proteins consists of five known members for which the functions are largely unknown. Here, we characterized mice lacking SALM1/LRFN2 (SALM1 KO) known to associate with NMDA receptors (NMDARs) and found that these mice showed altered NMDAR-dependent synaptic transmission and plasticity, as expected, but unexpectedly also exhibited suppressed inhibitory synapse development and synaptic transmission. Behaviorally, SALM1 KO pups showed suppressed ultrasonic vocalization upon separation from their mothers and SALM1 KO adults showed enhanced responses to loud acoustic stimuli. These results suggest that SALM1/LRFN2 regulates excitatory synapse function, inhibitory synapse development, social communication, and acoustic startle behavior.
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http://dx.doi.org/10.1523/JNEUROSCI.3321-17.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6595971PMC
June 2018

Bony cochlear nerve canal stenosis in pediatric unilateral sensorineural hearing loss.

Int J Pediatr Otorhinolaryngol 2018 Mar 2;106:72-74. Epub 2018 Feb 2.

Department of Otolaryngology, Daejeon St. Mary's Hospital, College of Medicine, The Catholic University of Korea, South Korea. Electronic address:

Objectives: This study was performed to evaluate the frequency of bony cochlear nerve canal (BCNC) stenosis and its clinical significance in pediatric patients with unilateral sensorineural hearing loss (SNHL) of unknown etiology.

Materials And Methods: We analyzed the medical records and temporal bone computed tomography (CT) results of patients less than 13 years of age with a diagnosis of unilateral SNHL of unknown etiology between July 2007 and July 2017. We compared the BCNC diameter between both sides and analyzed the age at diagnosis, degree of hearing loss, and accompanying inner ear anomalies.

Results: In 42 patients, the mean age at diagnosis was 7.4 ± 3.6 years, and the average hearing level in the affected ear was 87.9 ± 20.0 dB HL (decibels hearing level). The average diameter of the BCNC was 1.22 ± 0.75 mm on the affected side and 1.96 ± 0.52 mm on the normal side. The most suitable criterion for BCNC stenosis appeared to be a diameter of 1.2 mm by the recursive partitioning procedure. With application of this criterion, the rate of BCNC stenosis was significantly greater on the affected side than on the normal side (52.4% vs. 4.8%, respectively; P < 0.05). A narrow internal acoustic canal was found in two patients, and vestibular and cochlear anomalies were found in three patients each.

Conclusions: Our results suggest that it is reasonable to set a diameter of 1.2 mm as a cutoff for BCNC stenosis, and also that BCNC stenosis is a common cause of unilateral SNHL of unknown etiology in childhood.
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http://dx.doi.org/10.1016/j.ijporl.2018.01.015DOI Listing
March 2018

LAR-RPTP Clustering Is Modulated by Competitive Binding between Synaptic Adhesion Partners and Heparan Sulfate.

Front Mol Neurosci 2017 13;10:327. Epub 2017 Oct 13.

Center for Synaptic Brain Dysfunctions, Institute for Basic Science (IBS), Daejeon, South Korea.

The leukocyte common antigen-related receptor protein tyrosine phosphatases (LAR-RPTPs) are cellular receptors of heparan sulfate (HS) and chondroitin sulfate (CS) proteoglycans that direct axonal growth and neuronal regeneration. LAR-RPTPs are also synaptic adhesion molecules that form -synaptic adhesion complexes by binding to various postsynaptic adhesion ligands, such as Slit- and Trk-like family of proteins (Slitrks), IL-1 receptor accessory protein-like 1 (IL1RAPL1), interleukin-1 receptor accessory protein (IL-1RAcP) and neurotrophin receptor tyrosine kinase C (TrkC), to regulate synaptogenesis. Here, we determined the crystal structure of the human LAR-RPTP/IL1RAPL1 complex and found that lateral interactions between neighboring LAR-RPTP/IL1RAPL1 complexes in crystal lattices are critical for the higher-order assembly and synaptogenic activity of these complexes. Moreover, we found that LAR-RPTP binding to the postsynaptic adhesion ligands, Slitrk3, IL1RAPL1 and IL-1RAcP, but not TrkC, induces reciprocal higher-order clustering of -synaptic adhesion complexes. Although LAR-RPTP clustering was induced by either HS or postsynaptic adhesion ligands, the dominant binding of HS to the LAR-RPTP was capable of dismantling pre-established LAR-RPTP-mediated -synaptic adhesion complexes. These findings collectively suggest that LAR-RPTP clustering for synaptogenesis is modulated by a complex synapse-organizing protein network.
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http://dx.doi.org/10.3389/fnmol.2017.00327DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5645493PMC
October 2017

Autosomal dominant transmission of complicated hereditary spastic paraplegia due to a dominant negative mutation of KIF1A, SPG30 gene.

Sci Rep 2017 10 2;7(1):12527. Epub 2017 Oct 2.

Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea.

KIF1A is a brain-specific anterograde motor protein that transports cargoes towards the plus-ends of microtubules. Many variants of the KIF1A gene have been associated with neurodegenerative diseases and developmental delay. Homozygous mutations of KIF1A have been identified in a recessive subtype of hereditary spastic paraplegia (HSP), SPG30. In addition, KIF1A mutations have been found in pure HSP with autosomal dominant inheritance. Here we report the first case of familial complicated HSP with a KIF1A mutation transmitted in autosomal dominant inheritance. A heterozygous p.T258M mutation in KIF1A was found in a Korean family through targeted exome sequencing. They displayed phenotypes of mild intellectual disability with language delay, epilepsy, optic nerve atrophy, thinning of corpus callosum, periventricular white matter lesion, and microcephaly. A structural modeling revealed that the p.T258M mutation disrupted the binding of KIF1A motor domain to microtubules and its movement along microtubules. Assays of peripheral accumulation and proximal distribution of KIF1A motor indicated that the KIF1A motor domain with p.T258M mutation has reduced motor activity and exerts a dominant negative effect on wild-type KIF1A. These results suggest that the p.T258M mutation suppresses KIF1A motor activity and induces complicated HSP accompanying intellectual disability transmitted in autosomal dominant inheritance.
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http://dx.doi.org/10.1038/s41598-017-12999-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5624960PMC
October 2017

Phosphorylation of CYFIP2, a component of the WAVE-regulatory complex, regulates dendritic spine density and neurite outgrowth in cultured hippocampal neurons potentially by affecting the complex assembly.

Neuroreport 2017 Aug;28(12):749-754

Departments of aNeurosciencebBiomedical SciencescAnatomy, College of Medicine, Korea University, SeouldCenter for Synaptic Brain Dysfunctions, Institute for Basic Science (IBS), Daejeon, South Korea.

Actin dynamics is a critical mechanism underlying many cellular processes in neurons. The heteropentameric WAVE-regulatory complex (WRC), consisting of WAVE, CYFIP1/2, Nap, Abi, and HSPC300, is a key regulator of actin dynamics that activates the Arp2/3 complex to initiate actin polymerization and branching. The WRC is basally inactive because of intermolecular interactions among the components, which can be modulated by bindings of phospholipids and Rac1, and phosphorylations of WAVE and Abi. However, the phosphorylation of other components of WRC and their functional significance remain largely unknown. To address this issue, we focused on CYFIP1/2, in which we found two brain-specific phosphorylation sites (S582 of CYFIP2 and T1068/T1067 of CYFIP1/2) from a publicly available phosphoproteome database. To understand their functional effects, we overexpressed wild-type, phospho-blocking, or phospho-mimetic mutants of CYFIP2 in cultured hippocampal neurons, and found that only T1067A CYFIP2 decreased the density of stubby spines. Moreover, overexpression of wild-type CYFIP2 increased neurite length, but T1067A did not exert this effect. To understand the mechanism, we modeled CYFIP2 phosphorylation in the crystal structure of WRC and found that T1067 phosphorylation could weaken the interaction between CYFIP2 and Nap1 by inducing conformational changes of CYFIP2 α-helical bundles. In the co-immunoprecipitation assay, however, wild-type, T1067A, and T1067E CYFIP2 showed similar interaction levels to Nap1, suggesting that T1067 phosphorylation alone is not sufficient to disrupt the interaction. Considering that the activation of WRC requires disassembly of the complex, our results suggest that T1067 phosphorylation, together with other factors, could contribute toward the activation process.
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http://dx.doi.org/10.1097/WNR.0000000000000838DOI Listing
August 2017

Structural Insights into Modulation of Neurexin-Neuroligin Trans-synaptic Adhesion by MDGA1/Neuroligin-2 Complex.

Neuron 2017 Jun;94(6):1121-1131.e6

Center for Synaptic Brain Dysfunctions, Institute for Basic Science, Daejeon 34141, Korea; Graduate School of Medical Science & Engineering, KAIST, Daejeon 34141, Korea. Electronic address:

Membrane-associated mucin domain-containing glycosylphosphatidylinositol anchor proteins (MDGAs) bind directly to neuroligin-1 (NL1) and neuroligin-2 (NL2), thereby respectively regulating excitatory and inhibitory synapse development. However, the mechanisms by which MDGAs modulate NL activity to specify development of the two synapse types remain unclear. Here, we determined the crystal structures of human NL2/MDGA1 Ig1-3 complex, revealing their stable 2:2 arrangement with three interaction interfaces. Cell-based assays using structure-guided, site-directed MDGA1 mutants showed that all three contact patches were required for the MDGA's negative regulation of NL2-mediated synaptogenic activity. Furthermore, MDGA1 competed with neurexins for NL2 via its Ig1 domain. The binding affinities of both MDGA1 and MDGA2 for NL1 and NL2 were similar, consistent with the structural prediction of similar binding interfaces. However, MDGA1 selectively associated with NL2, but not NL1, in vivo. These findings collectively provide structural insights into the mechanism by which MDGAs negatively modulate synapse development governed by NLs/neurexins.
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http://dx.doi.org/10.1016/j.neuron.2017.05.034DOI Listing
June 2017

Structural and functional study of ChuY from Escherichia coli strain CFT073.

Biochem Biophys Res Commun 2017 Jan 3;482(4):1176-1182. Epub 2016 Dec 3.

Department of Molecular Cell Biology, Sungkyunkwan University School of Medicine, Suwon, Gyeonggi 16419, South Korea. Electronic address:

The uropathogenic Escherichia coli strain CFT073 contains multiple iron and heme transport systems, which facilitate infection of the host urinary tract. To elucidate the molecular and cellular function of ChuY, a hypothetical gene in the heme degradation/utilization pathway, we solved the crystal structure of ChuY at 2.4 Å resolution. ChuY has high structural homology with human biliverdin and flavin reductase. We confirmed that ChuY has flavin mononucleotide (FMN) reductase activity, using NAD(P)H as a cofactor, and shows porphyrin ring binding affinity. A chuY deletion-insertion strain showed reduced survival potential compared to wild-type and complemented strains in mammalian cells. Current results suggest ChuY acts as a reductase in heme homeostasis to maintain the virulence potential of E. coli CFT073.
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http://dx.doi.org/10.1016/j.bbrc.2016.12.008DOI Listing
January 2017

SALM4 suppresses excitatory synapse development by cis-inhibiting trans-synaptic SALM3-LAR adhesion.

Nat Commun 2016 08 2;7:12328. Epub 2016 Aug 2.

Department of Biological Sciences, Korea Advanced Institute for Science and Technology, Daejeon 305-701, Korea.

Synaptic adhesion molecules regulate various aspects of synapse development, function and plasticity. These functions mainly involve trans-synaptic interactions and positive regulations, whereas cis-interactions and negative regulation are less understood. Here we report that SALM4, a member of the SALM/Lrfn family of synaptic adhesion molecules, suppresses excitatory synapse development through cis inhibition of SALM3, another SALM family protein with synaptogenic activity. Salm4-mutant (Salm4(-/-)) mice show increased excitatory synapse numbers in the hippocampus. SALM4 cis-interacts with SALM3, inhibits trans-synaptic SALM3 interaction with presynaptic LAR family receptor tyrosine phosphatases and suppresses SALM3-dependent presynaptic differentiation. Importantly, deletion of Salm3 in Salm4(-/-) mice (Salm3(-/-); Salm4(-/-)) normalizes the increased excitatory synapse number. These results suggest that SALM4 negatively regulates excitatory synapses via cis inhibition of the trans-synaptic SALM3-LAR adhesion.
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http://dx.doi.org/10.1038/ncomms12328DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4974644PMC
August 2016

Structural and functional studies of a large winged Z-DNA-binding domain of Danio rerio protein kinase PKZ.

FEBS Lett 2016 07 23;590(14):2275-85. Epub 2016 Jun 23.

Department of Molecular Cell Biology, Sungkyunkwan University School of Medicine, Suwon, Korea.

The Z-DNA-binding domain of PKZ from zebrafish (Danio rerio; drZαPKZ ) contains the largest β-wing among known Z-DNA-binding domains. To elucidate the functional implication of the β-wing, we solved the crystal structure of apo-drZαPKZ . Structural comparison with its Z-DNA-bound form revealed a large conformational change within the β-wing during Z-DNA binding. Biochemical studies of protein mutants revealed that two basic residues in the β-wing are responsible for Z-DNA recognition as well as fast B-Z transition. Therefore, the extra basic residues in the β-wing of drZαPKZ are necessary for the fast B-Z transition activity.
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http://dx.doi.org/10.1002/1873-3468.12238DOI Listing
July 2016

SALM5 trans-synaptically interacts with LAR-RPTPs in a splicing-dependent manner to regulate synapse development.

Sci Rep 2016 05 26;6:26676. Epub 2016 May 26.

Department of Biological Sciences, Korea Advanced Institute for Science and Technology (KAIST), Daejeon 305-701, Korea.

Synaptogenic adhesion molecules play critical roles in synapse formation. SALM5/Lrfn5, a SALM/Lrfn family adhesion molecule implicated in autism spectrum disorders (ASDs) and schizophrenia, induces presynaptic differentiation in contacting axons, but its presynaptic ligand remains unknown. We found that SALM5 interacts with the Ig domains of LAR family receptor protein tyrosine phosphatases (LAR-RPTPs; LAR, PTPδ, and PTPσ). These interactions are strongly inhibited by the splice insert B in the Ig domain region of LAR-RPTPs, and mediate SALM5-dependent presynaptic differentiation in contacting axons. In addition, SALM5 regulates AMPA receptor-mediated synaptic transmission through mechanisms involving the interaction of postsynaptic SALM5 with presynaptic LAR-RPTPs. These results suggest that postsynaptic SALM5 promotes synapse development by trans-synaptically interacting with presynaptic LAR-RPTPs and is important for the regulation of excitatory synaptic strength.
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http://dx.doi.org/10.1038/srep26676DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4881023PMC
May 2016

Photocurrent enhancement of SiNW-FETs by integrating protein-shelled CdSe quantum dots.

Nanoscale 2016 Jan;8(4):1921-5

Department of Molecular Cell Biology, Sungkyunkwan University School of Medicine, Suwon 440-746, South Korea. and Sungkyunkwan Advanced Institute of Nanotechnology (SAINT), Sungkyunkwan University, Suwon 440-746, South Korea.

We proposed a new strategy to increase the photoresponsivity of silicon NW field-effect transistors (FETs) by integrating CdSe quantum dots (QDs) using protein shells (PSs). CdSe QDs were synthesized using ClpP, a bacterial protease, as protein shells to control the size and stability of QD and to facilitate the mounting of QDs on SiNWs. The photocurrent of SiNW-FETs in response to light at a wavelength of 480 nm was enhanced by a factor of 6.5 after integrating CdSe QDs because of the coupling of the optical properties of SiNWs and QDs. As a result, the photoresponsivity to 480 nm light reached up to 3.1 × 10(6), the highest value compared to other SiNW-based devices in the visible light range.
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http://dx.doi.org/10.1039/c5nr07901bDOI Listing
January 2016

Synaptic adhesion molecule IgSF11 regulates synaptic transmission and plasticity.

Nat Neurosci 2016 Jan 23;19(1):84-93. Epub 2015 Nov 23.

Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 305-701, Korea.

Synaptic adhesion molecules regulate synapse development and plasticity through mechanisms that include trans-synaptic adhesion and recruitment of diverse synaptic proteins. We found that the immunoglobulin superfamily member 11 (IgSF11), a homophilic adhesion molecule that preferentially expressed in the brain, is a dual-binding partner of the postsynaptic scaffolding protein PSD-95 and AMPA glutamate receptors (AMPARs). IgSF11 required PSD-95 binding for its excitatory synaptic localization. In addition, IgSF11 stabilized synaptic AMPARs, as determined by IgSF11 knockdown-induced suppression of AMPAR-mediated synaptic transmission and increased surface mobility of AMPARs, measured by high-throughput, single-molecule tracking. IgSF11 deletion in mice led to the suppression of AMPAR-mediated synaptic transmission in the dentate gyrus and long-term potentiation in the CA1 region of the hippocampus. IgSF11 did not regulate the functional characteristics of AMPARs, including desensitization, deactivation or recovery. These results suggest that IgSF11 regulates excitatory synaptic transmission and plasticity through its tripartite interactions with PSD-95 and AMPARs.
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http://dx.doi.org/10.1038/nn.4176DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5010778PMC
January 2016

Crystal structure analysis of c4763, a uropathogenic Escherichia coli-specific protein.

Acta Crystallogr F Struct Biol Commun 2015 Aug 29;71(Pt 8):1042-7. Epub 2015 Jul 29.

Department of Molecular Cell Biology, Sungkyunkwan University School of Medicine, Suwon 440-746, Republic of Korea.

Urinary-tract infections (UTIs), which are some of the most common infectious diseases in humans, can cause sepsis and death without proper treatment. Therefore, it is necessary to understand their pathogenicity for proper diagnosis and therapeutics. Uropathogenic Escherichia coli, the major causative agents of UTIs, contain several genes that are absent in nonpathogenic strains and are therefore considered to be relevant to UTI pathogenicity. c4763 is one of the uropathogenic E. coli-specific proteins, but its function is unknown. To investigate the function of c4763 and its possible role in UTI pathogenicity, its crystal structure was determined at a resolution of 1.45 Å by a multiple-wavelength anomalous diffraction method. c4763 is a homodimer with 129 residues in one subunit that contains a GGCT-like domain with five α-helices and seven β-strands. c4763 shows structural similarity to the C-terminal domain of allophanate hydrolase from Kluyveromyces lactis, which is involved in the degradation of urea. These results suggest that c4763 might be involved in the utilization of urea, which is necessary for bacterial survival in the urinary tract. Further biochemical and physiological investigation will elucidate its functional relevance in UTIs.
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http://dx.doi.org/10.1107/S2053230X15013035DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4528939PMC
August 2015

Dominant transmission of de novo KIF1A motor domain variant underlying pure spastic paraplegia.

Eur J Hum Genet 2015 Oct 14;23(10):1427-30. Epub 2015 Jan 14.

Research Programs Unit, Molecular Neurology, Biomedicum Helsinki, University of Helsinki, Helsinki, Finland.

Variants in family 1 kinesin (KIF1A), which encodes a kinesin axonal motor protein, have been described to cause variable neurological manifestations. Recessive missense variants have led to spastic paraplegia, and recessive truncations to sensory and autonomic neuropathy. De novo missense variants cause developmental delay or intellectual disability, cerebellar atrophy and variable spasticity. We describe a family with father-to-son transmission of de novo variant in the KIF1A motor domain, in a phenotype of pure spastic paraplegia. Structural modeling of the predicted p.(Ser69Leu) amino acid change suggested that it impairs the stable binding of ATP to the KIF1A protein. Our study reports the first dominantly inherited KIF1A variant and expands the spectrum of phenotypes caused by heterozygous KIF1A motor domain variants to include pure spastic paraplegia. We conclude that KIF1A should be considered a candidate gene for hereditary paraplegias regardless of inheritance pattern.
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http://dx.doi.org/10.1038/ejhg.2014.297DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4592090PMC
October 2015

Structural basis for LAR-RPTP/Slitrk complex-mediated synaptic adhesion.

Nat Commun 2014 Nov 14;5:5423. Epub 2014 Nov 14.

Graduate School of Medical Science and Engineering, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 305-701, Korea.

Synaptic adhesion molecules orchestrate synaptogenesis. The presynaptic leukocyte common antigen-related receptor protein tyrosine phosphatases (LAR-RPTPs) regulate synapse development by interacting with postsynaptic Slit- and Trk-like family proteins (Slitrks), which harbour two extracellular leucine-rich repeats (LRR1 and LRR2). Here we identify the minimal regions of the LAR-RPTPs and Slitrks, LAR-RPTPs Ig1-3 and Slitrks LRR1, for their interaction and synaptogenic function. Subsequent crystallographic and structure-guided functional analyses reveal that the splicing inserts in LAR-RPTPs are key molecular determinants for Slitrk binding and synapse formation. Moreover, structural comparison of the two Slitrk1 LRRs reveal that unique properties on the concave surface of Slitrk1 LRR1 render its specific binding to LAR-RPTPs. Finally, we demonstrate that lateral interactions between adjacent trans-synaptic LAR-RPTPs/Slitrks complexes observed in crystal lattices are critical for Slitrk1-induced lateral assembly and synaptogenic activity. Thus, we propose a model in which Slitrks mediate synaptogenic functions through direct binding to LAR-RPTPs and the subsequent lateral assembly of LAR-RPTPs/Slitrks complexes.
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http://dx.doi.org/10.1038/ncomms6423DOI Listing
November 2014

De novo mutations in the motor domain of KIF1A cause cognitive impairment, spastic paraparesis, axonal neuropathy, and cerebellar atrophy.

Hum Mutat 2015 Jan 27;36(1):69-78. Epub 2014 Nov 27.

Biomedical Proteomics Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea.

KIF1A is a neuron-specific motor protein that plays important roles in cargo transport along neurites. Recessive mutations in KIF1A were previously described in families with spastic paraparesis or sensory and autonomic neuropathy type-2. Here, we report 11 heterozygous de novo missense mutations (p.S58L, p.T99M, p.G102D, p.V144F, p.R167C, p.A202P, p.S215R, p.R216P, p.L249Q, p.E253K, and p.R316W) in KIF1A in 14 individuals, including two monozygotic twins. Two mutations (p.T99M and p.E253K) were recurrent, each being found in unrelated cases. All these de novo mutations are located in the motor domain (MD) of KIF1A. Structural modeling revealed that they alter conserved residues that are critical for the structure and function of the MD. Transfection studies suggested that at least five of these mutations affect the transport of the MD along axons. Individuals with de novo mutations in KIF1A display a phenotype characterized by cognitive impairment and variable presence of cerebellar atrophy, spastic paraparesis, optic nerve atrophy, peripheral neuropathy, and epilepsy. Our findings thus indicate that de novo missense mutations in the MD of KIF1A cause a phenotype that overlaps with, while being more severe, than that associated with recessive mutations in the same gene.
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http://dx.doi.org/10.1002/humu.22709DOI Listing
January 2015

Structural and kinetic bases for the metal preference of the M18 aminopeptidase from Pseudomonas aeruginosa.

Biochem Biophys Res Commun 2014 Apr 1;447(1):101-7. Epub 2014 Apr 1.

Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon 440-746, Republic of Korea. Electronic address:

The peptidases in clan MH are known as cocatalytic zinc peptidases that have two zinc ions in the active site, but their metal preference has not been rigorously investigated. In this study, the molecular basis for metal preference is provided from the structural and biochemical analyses. Kinetic studies of Pseudomonas aeruginosa aspartyl aminopeptidase (PaAP) which belongs to peptidase family M18 in clan MH revealed that its peptidase activity is dependent on Co(2+) rather than Zn(2+): the kcat (s(-1)) values of PaAP were 0.006, 5.10 and 0.43 in no-metal, Co(2+), and Zn(2+)conditions, respectively. Consistently, addition of low concentrations of Co(2+) to PaAP previously saturated with Zn(2+) greatly enhanced the enzymatic activity, suggesting that Co(2+)may be the physiologically relevant cocatalytic metal ion of PaAP. The crystal structures of PaAP complexes with Co(2+) or Zn(2+) commonly showed two metal ions in the active site coordinated with three conserved residues and a bicarbonate ion in a tetragonal geometry. However, Co(2+)- and Zn(2+)-bound structures showed no noticeable alterations relevant to differential effects of metal species, except the relative orientation of Glu-265, a general base in the active site. The characterization of mutant PaAP revealed that the first metal binding site is primarily responsible for metal preference. Similar to PaAP, Streptococcus pneumonia glutamyl aminopeptidase (SpGP), belonging to aminopeptidase family M42 in clan MH, also showed requirement for Co(2+) for maximum activity. These results proposed that clan MH peptidases might be a cocatalytic cobalt peptidase rather than a zinc-dependent peptidase.
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http://dx.doi.org/10.1016/j.bbrc.2014.03.109DOI Listing
April 2014

Distinct Rayleigh scattering from hot spot mutant p53 proteins reveals cancer cells.

Small 2014 Jul 3;10(14):2954-62. Epub 2014 Apr 3.

Department of Chemical and Biological Engineering, Korea University, Seoul, 136-701, Korea.

The scattering of light redirects and resonances when an electromagnetic wave interacts with electrons orbits in the hot spot core protein and oscillated electron of the gold nanoparticles (AuNP). This report demonstrates convincingly that resonant Rayleigh scattering generated from hot spot mutant p53 proteins is correspondence to cancer cells. Hot spot mutants have unique local electron density changes that affect specificity of DNA binding affinity compared with wild types. Rayleigh scattering changes introduced by hot-spot mutations were monitored by localized surface plasmon resonance (LSPR) shift changes. The LSPR λmax shift for hot-spot mutants ranged from 1.7 to 4.2 nm for mouse samples and from 0.64 nm to 2.66 nm for human samples, compared to 9.6 nm and 15 nm for wild type and mouse and human proteins, respectively with a detection sensitivity of p53 concentration at 17.9 nM. It is interesting that hot-spot mutants, which affect only interaction with DNA, launches affinitive changes as considerable as wild types. These changes propose that hot-spot mutants p53 proteins can be easily detected by local electron density alterations that disturbs the specificity of DNA binding of p53 core domain on the surface of the DNA probed-nanoplasmonic sensor.
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http://dx.doi.org/10.1002/smll.201400004DOI Listing
July 2014