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Production and structural characterization of human Gla-rich Protein, an inhibitor of vascular calcification
ID Janakievski, Nikola (Avtor), ID Gaber, Aljaž (Mentor) Več o mentorju... Povezava se odpre v novem oknu, ID dos Anjos López de Macedo, Maria (Komentor)

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Izvleček
Gla-rich protein (GRP) is a novel protein connected to multiple pathological disorders plaguing humanity in modern times, including chronic kidney disease, atherosclerosis, osteoarthritis, and cancer. It gets its name from the high number of γ-carboxylated glutamate residues, most of any known protein, this enables it to have a very high affinity for calcium ions, making it a potential inhibitor of ectopic calcification in soft tissue. Except for the fact that this protein contains intrinsically disordered domains, not much else is known about the structure of GRP. NMR spectroscopy is the perfect structure determination technique to tackle this problem, specializing in the structural determination of small proteins and more recently for intrinsically disordered proteins. We expressed the human GRP variant in transformed E. coli cells and labelled it with isotopes 13C and 15N. After purification, we prepared samples for liquid-state NMR spectroscopy and explored the most practical NMR experiments for structure determination. To conclude, we found the optimal experimental conditions for NMR sample preparation of human GRP allowing for structural studies to be performed. We tested out different 15N and 13C direct detection experiments at different acquisition temperatures. Our preliminary data is promising and with the acquisition of other types of multidimensional NMR spectra, and after resonance assignment, structural information can be obtained for human GRP structural features elucidation.

Jezik:Angleški jezik
Ključne besede:NMR, ectopic calcification, human GRP, expression, IDPs
Vrsta gradiva:Diplomsko delo/naloga
Tipologija:2.11 - Diplomsko delo
Organizacija:FKKT - Fakulteta za kemijo in kemijsko tehnologijo
Leto izida:2022
PID:20.500.12556/RUL-143282 Povezava se odpre v novem oknu
COBISS.SI-ID:138709763 Povezava se odpre v novem oknu
Datum objave v RUL:12.12.2022
Število ogledov:647
Število prenosov:46
Metapodatki:XML RDF-CHPDL DC-XML DC-RDF
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Sekundarni jezik

Jezik:Slovenski jezik
Naslov:Priprava in strukturna karakterizacija inhibitorja žilne kalcinacije, človeškega s karboksiglutaminsko kislino bogatega proteina (GRP)
Izvleček:
Gla-rich protein (GRP) is a novel protein connected to multiple pathological disorders plaguing humanity in modern times, including chronic kidney disease, atherosclerosis, osteoarthritis, and cancer. It gets its name from the high number of γ-carboxylated glutamate residues, most of any known protein, this enables it to have a very high affinity for calcium ions, making it a potential inhibitor of ectopic calcification in soft tissue. Except for the fact that this protein contains intrinsically disordered domains, not much else is known about the structure of GRP. NMR spectroscopy is the perfect structure determination technique to tackle this problem, specializing in the structural determination of small proteins and more recently for intrinsically disordered proteins. We expressed the human GRP variant in transformed E. coli cells and labelled it with isotopes 13C and 15N. After purification, we prepared samples for liquid-state NMR spectroscopy and explored the most practical NMR experiments for structure determination. To conclude, we found the optimal experimental conditions for NMR sample preparation of human GRP allowing for structural studies to be performed. We tested out different 15N and 13C direct detection experiments at different acquisition temperatures. Our preliminary data is promising and with the acquisition of other types of multidimensional NMR spectra, and after resonance assignment, structural information can be obtained for human GRP structural features elucidation.

Ključne besede:NMR, ektopična kalcifikacija, človeški GRP, ekspresija, IDP

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