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Cooperability
| Project leader |
Husnjak Koraljka |
| Project co-leader: |
Mihaela Matovina |
| Administering organization: |
Ruđer Bošković Institute, Bijenička cesta 54, HR-10000 Zagreb, Croatia |
| Partner Institution/Company: |
Institute of Biochemistry II, Goethe University School of Medicine, Frankfurt am Main |
| Grant type: |
1B |
| Project title: |
Elucidation of the physiological roles of human dipeptidyl peptidase III |
| Project summary: |
Proteolytic enzymes (peptidases) participate in and regulate an increasing number of important cellular processes by specifically cleaving their target proteins. Although structurally and biochemically very well characterized, many peptidases still lack information about their specific substrates and physiological roles.
Dipeptidyl peptidase III (DPP III) is the cytosolic metallopeptidase that is well characterized at the structural and enzymatic level, however its physiological roles are still largely unknown. DPP III peptidase activity towards four to ten amino acid long oligopeptides indicates its role in the terminal stages of protein turnover, while its activity, and affinity towards certain biologically active peptides indicates its role in the pain modulation, blood pressure control, and inflammation. Furthermore, its involvement in the oxidative stress response through the interaction with Keap1 protein has been well documented. However, none of the proposed roles of DPP III have been proven, yet, and there are indications that it might also have other roles in the cell physiology. Its proposed role in Nrf2/Keap1 pathway, taken together with its overexpression in tumor tissues of several different tumor types, indicate that it might be a biomarker of certain types of cancer or even the target for anticancer therapy. The aim of this project is to elucidate the physiological roles, and disease relevance of DPP III by determining its interactors through the combined use of different molecular biology approaches. We plan to use yeast two-hybrid method to identify novel interacting partners of DPP III, and mass spectrometry interactome analysis to identify protein complexes DPP III takes part in. We plan to confirm the interactions found by Y2H and MS approach by several low-throughput methods. Finding DPP III interactors would enable us to identify other signaling pathways DPP III might be involved in. We will also test if any of newly identified DPP III interacting proteins are also DPP III substrates. Apart from gaining new insights into physiology of DPP III, this project will also enable transfer of technology and knowledge to the Ruđer Bošković Institute, which will have a positive impact on our future research efforts, and represent the first step towards procuring continuous research funding from EU funds.
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| Hrvatski sažetak: |
Proteolitički enzimi (peptidaze) sudjeluju u regulaciji velikog broja ključnih procesa u stanici specifičnim cijepanjem odabranih ciljnih proteina. Iako su strukturno i biokemijski detaljno okarakterizirane, za mnoge peptidaze još uvijek nedostaju podaci o njihovim specifičnim supstratima i fiziološkoj ulozi. Dipeptidil peptidaza III (DPP III) je citoplazmatska metalopeptidaza detaljno okarakterizirana na strukturnoj i enzimatskoj razini, ali njezina fiziološka uloga još je gotovo potpuno nepoznata. Aktivnost DPP III peptidaze prema četiri do deset aminokiselina dugim peptidima sugerira njenu ulogu u završnim fazama razgradnje proteina, dok njezina aktivnost i afinitet prema nekim biološki aktivnim peptidima upućuje na njenu ulogu u regulaciji boli, krvnog tlaka i upali. Nadalje, uključenost DPP III u odgovor na oksidativni stres putem interakcije s proteinom Keap1 detaljno je opisana, međutim niti jedna od predloženih funkcija proteina DPP III još nije dokazana, a također postoje indikacije da bi mogao imati ulogu i u drugim fiziološkim procesima u stanici. Predložena funkcija u signalnom putu Nrf2/Keap1, kao i česta prekomjerna ekspresija proteina DPP III u različitim tipovima tumora, sugeriraju da bi mogao biti korišten kao tumorski biljeg ili čak meta antitumorske terapije.
Cilj ovog projekta je određivanje fiziološke uloge i važnosti proteina DPP III u bolesti određivanjem njegovih interakcijskih proteina pomoću različitih molekularno-bioloških metoda. U radu ćemo koristiti tzv. metodu "yeast two-hybrid" (Y2H) za identifikaciju novih interakcijskih partnera proteina DPP III, kao i masenu spektrometriju (MS), za identifikaciju proteinskih kompleksa koji uključuju DPP III. Namjeravamo potvrditi interakcijske partnere proteina DPP III dobivene metodom Y2H i MS korištenjem nekoliko dodatnih metoda. Potvrđeni interakcijski partneri pomoći će nam u identifikaciji signalnih puteva koji uključuju DPP III. Također ćemo testirati da li su novoidentificirani interakcijski partneri proteina DPP III i njegovi supstrati. Osim dobivanja uvida u fiziologiju proteina DPP III, ovaj projekt omogućit će prijenos tehnologije i znanja u Institut Ruđer Bošković, što će pozitivno utjecati na naš budući znanstveni rad, kao i predstavljati prvi korak u osiguravanju trajne financijske potpore istraživanja iz europskih fondova.
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| Amount requested from UKF: |
1,409.411,76 |
| Amount of matching funding: |
295.470,59 |
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