Hydrophilic spacer-arm containing magnetic nanoparticles for immobilization of proteinase K: Employment for speciation of proteins for mass spectrometry-based analysis

Basit Öğe Kaydı
dc.contributor.authorBayramoglu, Gulay
dc.contributor.authorKayili, Haci Mehmet
dc.contributor.authorOztekin, Merve
dc.contributor.authorSalih, Bekir
dc.contributor.authorArica, M. Yakup
dc.date.accessioned2024-09-29T16:00:48Z
dc.date.available2024-09-29T16:00:48Z
dc.date.issued2020
dc.departmentKarabük Üniversitesien_US
dc.description.abstractProteinase K (ProK) is used for the degradation of proteins in cell lysates to isolate nucleic acids, and for the speciation of proteins for mass spectrometry analysis. In this work, a novel and sensitive immobilization process was developed for examination of protein mixtures by combining MALDI-ToF-MS and nLC-TIMS-ToF-MS/MS systems. To achieve these goals, magnetic nanoparticles (MPs) were prepared via thermal coprecipitation reaction under alkaline condition. The MPs were grafted with a silica layer (i.e., 3-(2,3-epoxypropoxy) propyltrimethoxysilane; EPTES) containing reactive epoxy groups. Then, the silica-grafted magnetic particles were coated with a long chain hydrophilic poly(ethylene glycol) diamine polymer (PEGDAP). The prepared materials were characterized by the Brunauer-Emmett-Teller (BET) method, X-ray diffraction (XRD), scanning electron microscopy (SEM) and attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy. The VSM data show that the MPs@EPTES@PEGDAP has paramagnetic performance with a saturation magnetization of approximately 32.3 emu g(-1). Proteinase K (EC 3.4.21.64) was covalently immobilized on the MPs@EPTES by reaction of its epoxy groups with amine groups of the enzyme. On the other hand, the ProK was immobilized on the MPs@EPTES@PEGDAP after activation with glutaraldehyde and the immobilization reaction was realized by the coupling reaction between aldehyde groups of the support and amine groups of the enzyme. The amounts of immobilized ProK on the MPs@EPTES and MPs@EPTES@PEGDAP were found to be 27.4 and 19.6 mg g(-1) land the retained activities were determined to be 29 and 87%, respectively. For the first time, some important features such as thermal and storage stabilities, reusability and potential use in protein speciation for mass spectrometry-based techniques were also evaluated. For examples, after six weeks of storage at 4 degrees C, the immobilized ProK on the MPs@EPTES@PEGDAP-ProK still maintained 59% of its initial activity. However, at the end of the six-week storage period, its free counterpart had lost all of its initial activity. The immobilized ProK was also utilized for degradation and identification of model proteins (i.e., alpha-2-HS glycoprotein, beta-casein, bovine serum albumin and immunoglobulin). After enzymatic treatment, the digested peptides were analyzed and mapped by using nLC-TIMS-ToF-MS/MS systems.en_US
dc.description.sponsorshipMinistry of Development-Republic of Turkey [FAY-2015-7201]en_US
dc.description.sponsorshipThis work was supported by Ministry of Development-Republic of Turkey with the project Number: FAY-2015-7201.en_US
dc.identifier.doi10.1016/j.talanta.2019.120218
dc.identifier.issn0039-9140
dc.identifier.issn1873-3573
dc.identifier.pmid31514867en_US
dc.identifier.scopus2-s2.0-85070239925en_US
dc.identifier.scopusqualityQ1en_US
dc.identifier.urihttps://doi.org/10.1016/j.talanta.2019.120218
dc.identifier.urihttps://hdl.handle.net/20.500.14619/5370
dc.identifier.volume206en_US
dc.identifier.wosWOS:000488416200034en_US
dc.identifier.wosqualityQ1en_US
dc.indekslendigikaynakWeb of Scienceen_US
dc.indekslendigikaynakScopusen_US
dc.indekslendigikaynakPubMeden_US
dc.language.isoenen_US
dc.publisherElsevieren_US
dc.relation.ispartofTalantaen_US
dc.relation.publicationcategoryMakale - Uluslararası Hakemli Dergi - Kurum Öğretim Elemanıen_US
dc.rightsinfo:eu-repo/semantics/closedAccessen_US
dc.subjectMagnetic nanoparticlesen_US
dc.subjectImmobilized proteinase Ken_US
dc.subjectPeptide synthesisen_US
dc.subjectProtein speciationen_US
dc.subjectMALDI-ToF-MSen_US
dc.titleHydrophilic spacer-arm containing magnetic nanoparticles for immobilization of proteinase K: Employment for speciation of proteins for mass spectrometry-based analysisen_US
dc.typeArticleen_US

Dosyalar

Koleksiyon

WoS İndeksli Yayınlar Koleksiyonu
PubMed İndeksli Yayınlar Koleksiyonu
Scopus İndeksli Yayınlar Koleksiyonu