Production Of EGFP Tagged Vm24 Peptide Toxin: A Fluorescent Probe Of Voltage-Gated Potassuim Channel Kv1.3

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dc.contributor.advisorPanyi, György
dc.contributor.advisordeptDepartment of Biophysics and Cell Biologyhu_HU
dc.contributor.authorAl Olaimi, Amna Sami
dc.contributor.departmentDE--Általános Orvostudományi Karhu_HU
dc.contributor.opponentKovács, Tünde
dc.contributor.opponentdeptOrvosi Vegytani Intézethu_HU
dc.date.accessioned2022-06-10T15:23:54Z
dc.date.available2022-06-10T15:23:54Z
dc.date.created2022-05-03
dc.description.abstractVenomous peptides identified in various animal species are being directed toward the treatment of T-cell mediated autoimmune disorders such as rheumatoid arthritis, systemic lupus erythematosus, and multiple sclerosis. The selectivity of the venomous peptides can be harnessed in the form of a molecular tool for various research applications by conjugating the peptide to trackable fluorescent proteins. A 36-residues long peptide toxin, Vm24, was purified from the venom of the scorpion Vaejovis mexicanus smithi which has a high affinity for Kv1.3 channels in human lymphocytes (Kd= 2.9 pM) and increased selectivity (>1500-fold) over other ion channels tested. In our approach, an N-terminal His6 tagged EGFP fluorescent protein is conjugated to Vm24 toxin and is expressed in Pichia pastoris. The Pichia clones were screened against a progressively increasing Zeocin™ concentration in search of hyper-resistant clone against Zeocin. and the Conjugate EGPF-Vm24 protein was purified using Ni+ affinity chromatography then further refined using reverse-phase HPLC. The activity of the conjugate was tested through electrophysiological recordings on activated human peripheral T lymphocytes using the whole-cell patch-clamp technique, the chimera protein retained the blocking activity towards Kv1.3 in picomolar range (Kd = 328 pM).hu_HU
dc.description.coursemolekuláris biológiahu_HU
dc.description.courselangangolhu_HU
dc.description.coursespecBiokémia-genomikahu_HU
dc.description.degreeMSc/MAhu_HU
dc.format.extent35hu_HU
dc.identifier.urihttp://hdl.handle.net/2437/334423
dc.language.isoenhu_HU
dc.subjectPotassuim channelshu_HU
dc.subjecttoxinshu_HU
dc.subjectvm24hu_HU
dc.subjectaffinity chromatographyhu_HU
dc.subjectEGFPhu_HU
dc.subjectRP-HPLChu_HU
dc.subjectPCRhu_HU
dc.subjectProtein purificationhu_HU
dc.subjectelectrophysiologyhu_HU
dc.subjectprotein productionhu_HU
dc.subject.dspaceDEENK Témalista::Orvostudományhu_HU
dc.titleProduction Of EGFP Tagged Vm24 Peptide Toxin: A Fluorescent Probe Of Voltage-Gated Potassuim Channel Kv1.3hu_HU
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