Design of a chromogenic substrate for elastase based on split GFP system proof of concept for colour switch sensors

التفاصيل البيبلوغرافية
العنوان: Design of a chromogenic substrate for elastase based on split GFP system proof of concept for colour switch sensors
المؤلفون: Ana Vanessa Fernandes Ferreira, Teresa Matamá, Nuno G. Azoia, Joana Cunha, Artur Cavaco-Paulo, Artur Ribeiro, Egipto Antunes
المساهمون: Universidade do Minho
المصدر: Repositório Científico de Acesso Aberto de Portugal
Repositório Científico de Acesso Aberto de Portugal (RCAAP)
instacron:RCAAP
Biotechnology Reports
Biotechnology Reports, Vol 22, Iss, Pp-(2019)
بيانات النشر: Elsevier BV, 2019.
سنة النشر: 2019
مصطلحات موضوعية: UM, ultramarine protein, UM10, ultramarine engineered without the last 11th β-strand, lcsh:Biotechnology, HNE, human neutrophil elastase, 010402 general chemistry, 01 natural sciences, Applied Microbiology and Biotechnology, Article, Green fluorescent protein, 03 medical and health sciences, Chromoprotein, lcsh:TP248.13-248.65, pep11, synthetic peptide equivalent to free 11th β-sheet of ultramarine, Moiety, E11, 11th β-strand of ultramarine inserted in a eglin c portion with an HNE cleavage sequence, 030304 developmental biology, ComputingMethodologies_COMPUTERGRAPHICS, High concentration, RMSF, root-mean-square fluctuation, 0303 health sciences, Chemistry, Chromogenic, Molecular dynamics simulations, Split GFP, Elastase, Ultramarine (UM), Substrate (chemistry), Chromogenic GFP-like proteins, Proteases, 0104 chemical sciences, Biophysics, Biosensor, pNa, p-nitroaniline, Human neutrophil elastase (HNE), Biotechnology
الوصف: Graphical abstract
Highlights • GFP-like chromoproteins can be used for protease signalling through colour changes. • Absorbance changes obtained on split UM system upon HNE cleavage were transient. • Molecular dynamics confirmed the instability registered in vitro studies. • In silico studies provide further understanding on split and reassembly mechanisms. • Split-GFP technology for chromoproteins isn’t straightforward as for fluoroproteins.
Recent studies have demonstrated that human neutrophil elastase (HNE) can be used as marker for inflammation/infection of chronic wounds since it was found to be present in high concentration in exudate collected from chronic wounds. Biosensors used in wound care benefit from a chromogenic signalling due to the readiness of signal interpretation, but the most common use faint yellow chromogenic molecules such as p-nitroaniline (pNa). In addition, if to be converted into smart dressings, the colour of the detection system should not be masked by the exudate’s colour. In this work, we designed a chromogenic substrate for HNE aiming to be incorporated in a smart dressing as a colour switch sensor. The substrate was developed using the GFP-like chromoprotein ultramarine (UM), following the split GFP technology. The cleavage sequence for HNE (Ala-Ala-Pro-Val) was embedded into the sensing moiety of the substrate corresponding to the 11th β-sheet. In the presence of HNE, the 11th β-sheet is able to interact to the signalling moiety composed of the β1-β10 incomplete barrel, allowing the re-establishment of the chromophore environment and, hence, the colour production. Structural homology and molecular dynamics simulations were conducted to aid on the disclosure of the structural changes that are the base of the mechanism of action of this HNE switch substrate. Our findings explore the possible application of GFP-like chromogenic sensors in point-of-care devices for the evaluation of the wounds status, representing a major step in the medical field.
وصف الملف: application/pdf
اللغة: English
URL الوصول: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::079d5d6d94984fcb8452e3dda46186e0
Rights: OPEN
رقم الانضمام: edsair.doi.dedup.....079d5d6d94984fcb8452e3dda46186e0
قاعدة البيانات: OpenAIRE