Študija zaviralcev amiloidnih bolezni z vibracijsko spektroskopijo

Kelhar, Tim (author), Zorko, Andrej (mentor), Novak, Urban (comentor)

Preučevali smo agregacijo konkanavalina A z uporabo infrardeče spektroskopije. Konkanavalin A je protein s pretežno sekundarno strukturo beta, ki ima zelo zanimivo vrsto agregacije. V kislem mediju se z nespremenjeno strukturo agregira v amorfne agregate, medtem ko v bazičnem okolju tvori amiloidne fibrile. Zato smo ga uporabili za testiranje zaviranja tvorbe amiloidov, ki so glavni povzročitelji tako imenovanih konformacijskih bolezni, kot sta npr. Parkinsonova in Alzheimerjeva bolezen. Tvorbo agregatov in amiloidov smo pospešili z dvigovanjem temperature. Za zaviranje smo uporabili dva znana zaviralca urejenih proteinskih agregatov, homotavrin in resveratrol. Izračunali smo sigmoidne krivulje ter določili in spremljali temperature prehoda iz raztopine v agregate, z in brez inhibitorja. Z metodo diferenčne spektroskopije smo spremljali majhne strukturne spremembe, ki nastanejo kot posledica zunanjih sprememb. Analizni spekter je izračunan kot razlika med dvema posnetima spektroma ob na primer različnih temperaturah. Zato nam ponuja metoda vpogled le v vrhove, ki niso skupni obema izhodnima spektroma. Temperatura prehoda ni naraščala, tako kot smo to pričakovali z naraščanjem koncentracije inhibitorja. Kjer se izkaže, da se je dvigovala, to ovržemo z negotovostjo meritev. Možnih razlogov za to je več, kriva je lahko napačna izbira koncentracijskih razmerij, nasičena raztopina, ali pa moja napaka pri mešanju raztopin. Največ težav pri raztapljanju smo imeli pri resveratrolu, zato mogoče ni bil najboljša izbira za izbrana koncentracijska razmerja.

• Language: Slovenian
• Keywords: protein, sekundarna struktura, konkanavalin A, agregacija, infrardeča spektroskopija, diferenčna spektroskopija, homotavrin, resveratrol
• Work type: Final paper
• Typology: 2.11 - Undergraduate Thesis
• Organization: FMF - Faculty of Mathematics and Physics
• Year: 2020
• PID: 20.500.12556/RUL-120307
• COBISS.SI-ID: 30082819
• Publication date in RUL: 18.09.2020

KELHAR, Tim, 2020, Študija zaviralcev amiloidnih bolezni z vibracijsko spektroskopijo [online]. Bachelor’s thesis. [Accessed 18 May 2025]. Retrieved from: https://repozitorij.uni-lj.si/IzpisGradiva.php?lang=eng&id=120307

Secondary language

• Language: English
• Title: Study of amyloid diseases by vibrational spectroscopy

Abstract

We investigated the aggregation of Concanavalin A using infrared spectroscopy. Concanavalin A is a protein with a predominantly $\beta$-structure and has a very interesting way of aggregation. In acidic medium it aggregates in amorphous aggregates, whereas in alkaline medium it forms amyloid fibrils. Therefore, we have used it for testing retarding amyloids, which are the main cause of conformation diseases, such as Parkinson and Alzheimer disease. The growth of aggregates and amyloids was accelerated with the increase in temperature. We used two well-known inhibitors to slow down settled protein aggregates, which are homotavrine and resveratrol. We calculate sigmoid curves from structural changes taken from the analysis of amide I band and monitored the temperature of the transition from solution to aggregates, with and without inhibitors. Using the method of differential spectroscopy, we were able to accompany small structural changes, which are caused by temperature changes. The analysis spectrum is calculated as the difference between two spectra recorded at different temperatures. For this reason, this method allows us to look into spectral bands that are not the same for both spectra. The temperature of the transition did not rise, as we had expected, with the increase in inhibitor concentration. Where it shows the increase, it correlates with the measurement error. There can be many reasons for this, it could be a wrong choice of concentration ratio, a saturated solution or my error with the solutions. Most of the problems with solving occurred with resveratrol, maybe it was not the best choice for the chosen concentration ratios.

• Keywords: protein, secondary structure, concanavalin A, agreggation, infrared spectroscopy, differential spectroscopy