Uloga p97/VCP AAA ATPaze u popravku unakrsnog vezanja DNA i proteina

master's thesis

• Title (english): The role of p97/VCP AAA ATPase in the repair of DNA-protein crosslinks
• Author: Valentin Dakić
• Mentor: Marta Popović (mentor)
• Committee member: Morana Dulić (predsjednik povjerenstva)
• Committee member: Iva Juranović Cindrić (član povjerenstva)
• Committee member: Đani Škalamera (član povjerenstva)
• Committee member: Marko Močibob (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Chemistry); Zagreb
• Defense date and country: 2020-12-22, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Chemistry

Abstract

Unakrsna veza između DNA i proteina (engl. DNA-protein crosslink, DPC) je tip oštećenja DNA koje nastaje kao posljedica stvaranja ireverzibilne kovalentne veze između DNA i proteina u prisutnosti endogenog ili egzogenog DPC-inducirajućeg agensa. Nemogućnost popravka DPC-a uzrokuje genomsku nestabilnost i teške posljedice za organizam, uključujući preuranjeno starenje, neurodegeneraciju i pojavu tumora. ATPaza p97 tvori kompleks s glavnim čimbenikom popravka DPC-a, SPRTN metaloproteazom. Međutim, uloga p97 u ovom putu DNA popravka nije poznata, te je stoga cilj ovog rada istražiti sudjeluje li i na koji način p97 u popravku DPC-a i u kakvom je odnosu sa SPRTN proteazom. U tu su svrhu prekomjerno eksprimirani i pročišćeni humani proteini p97 i njegov interaktor SPRTN radi kasnije strukturne i biokemijske analize njihovog kompleksa krio-elektronskom mikroskopijom. Provedena je i analiza ekspresije p97 i SPRTN gena (na razini mRNA) u tkivima dvaju modelnih organizama, ribe zebrice (Danio rerio) i domaćeg miša (Mus musculus), kako bi se usporedio ekspresijski profil p97 i SPRTN gena kroz tkiva te među organizmima.

Abstract (english)

DNA-protein crosslink (DPC) is a type of DNA damage that occurs as a result of the formation of an irreversible covalent bond between DNA and protein in the presence of an endogenous or exogenous DPC-inducing agent. The inability to repair DPCs causes genomic instability and severe consequences for the organism, including premature aging, neurodegeneration, and tumor appearance. ATPase p97 forms a complex with the major DPC repair factor, SPRTN metalloprotease. However, the role of p97 in this pathway of DNA repair is unknown, and therefore the aim of this thesis is to get closer to answering whether and in what way p97 participates in DPC repair and what is its relation to SPRTN protease. For this purpose, human proteins p97 and its interactor SPRTN were overexpressed and purified for subsequent structural and biochemical analysis of their complex by cryo-electron microscopy. Expression analysis of p97 and SPRTN genes (at the mRNA level) in the tissues of two model organisms, zebrafish (Danio rerio) and mice (Mus musculus) was performed to compare the expression profile of p97 and SPRTN genes across tissues and organisms.

• Keywords: ATPaza p97/VCP; ekspresija gena; izolacija proteina; izolacija ukupne RNA; kvantitativni PCR; metaloproteaza SPRTN; unakrsna veza DNA-protein
• Keywords (english): ATPase p97/VCP; DNA-protein crosslink; gene expression; protein isolation; quantitative PCR; SPRTN metalloprotease; total RNA isolation
• Language: croatian
• URN:NBN: urn:nbn:hr:217:726279
• Study programme: Title: Graduate university studies in chemistry; specializations in: Research programme; Course: Research programme; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra kemije (magistar/magistra kemije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access; Embargo expiration date: 2022-12-22
• Created on: 2021-02-16 12:23:14