Primjena antisens peptida kao novi pristup liječenja ugriza poskoka

master's thesis

• Title (english): Antisens peptides application as a new approach to horned viper envenomation treatment
• Author: Marin Marić
• Mentor: Mario Gabričević (mentor)
• Committee member: Mario Gabričević (predsjednik povjerenstva)
• Committee member: Tin Weitner (član povjerenstva)
• Committee member: Viktor Pilepić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Pharmacy and Biochemistry; (Department of general and inorganic chemistry); Zagreb
• Defense date and country: 2021-09-16, Croatia
• Scientific / art field, discipline and subdiscipline: BIOMEDICINE AND HEALTHCARE; Pharmacy; Pharmacy

Abstract

Danas se kao terapija ugriza poskoka koriste imunoglobulini zmijskog protuotrova čija je proizvodnja složena, skupa i
dugotrajna. Kao novi pristup liječenju predlaže se upotreba oligopeptida koji će se nekovalentno vezati na dijelove
amoditoksina A odgovorne za neurotoksičnost inhibirajući njegovo djelovanje. Dizajn takvih oligopeptida temelji se na
teoriji molekularnog prepoznavanja koja sugerira da svaka aminokiselina ulazi u interakciju s točno određenom
aminokiselinom. Cilj ovog istraživanja je odrediti afinitet vezanja sens peptida koji predstavljaju neurotoksične dijelove
amoditoksina A i njihovih potencijalno komplementarnih antisens peptida. Primarna struktura sens1 peptida je
GMMILG, a njegovih antisens peptida SHHYEP i SHHYSP. Primarna struktura sens2 peptida je YIYRNYPDF, a
njegovih antisens peptida INISIVGVK i VNISVIGIK. Konstante disocijacije određene su spektrofluorimetrijskim
određivanjem intenziteta fluorescencije tijekom titracije. Kao titranti su korišteni sens1, sens2antisens1 i sens2antisens2
peptidi jer ne fluoresciraju zbog odsustva aromatskih aminokiselina. Kao titrandi su korišteni sens1antisens1,
sens1antisens2 i sens2 peptidi koji fluoresciraju. Analizom eksperimentalno dobivenih podataka računalno su izračunate
konstante disocijacija. Njihova vrijednost je usporediva s vrijednostima konstantama disocijacije većine antitijela, što
upućuje na jaku interakciju, tj. afinitet vezanja. Kd za interakciju sens1 i njegovih antisens peptida su 5,071·10-6 M i
1,434·10-6 M, a za interakciju sens2 peptida i njegovih antisens peptida 1,143·10-6 M i 9,569·10-6 M. Navedeni rezultati
potvrđuju vezanje sens i antisens peptida i potencijalni terapijski učinak antisens peptida koji se mogu koristiti u daljnjim
istraživanjima. Pozitivan rezultat ovog istraživanja sugerira što veću upotrebu teorije molekularnog prepoznavanja za
dizajniranje antisens peptida koji bi se mogli koristiti u raznim područjima biomedicine.

Abstract (english)

Nowadays snake antivenom imunoglobulins are being used for snake bite treatment whose production is complex,
expensive and long. Use of oligopeptides is suggested as a new therapeutic approach which form noncovalent bonds
with ammodytoxin A neurotoxic sites inhibiting it's action. Oligopeptide design is based on molecular recognition theory
which indicates that every aminoacid can interact with the only specific complementary aminoacid. The aim of this
research is the determination of binding affinity of sens peptides which represent ammodytoxin A neurotoxic sites and
their potentially complementary antisens peptides. Primary structure of sens1 peptide is GMMILG and his antisens
peptides are SHHYEP and SHHYSP. Primary structure of sens2 peptide is YIYRNYPDF and his antisens peptides are
INISIVGVK and VNISVIGIK. Dissociation constants are determined by spectrofluorimetric measuring of fluorescence
intensity during titration. Sens1, sens2antisens1 and sens2antisens2 peptides were used as titrants considering the fact
they contain aromatic aminoacids which are responsible for fluorescence. On the other hand, sens1antisens1,
sens1antisens2 and sens2 peptides were used as titrands since they lack aromatic aminoacids. Computer program
analysed experimental data and calculated dissociaton constants. Their values are comparable to those of antibodies
which points to strong interaction, i. e. binding affinity. Kd values for sens1 peptide and it's antisens peptides interactions
are 5,071·10-6 M and 1,434·10-6 M and for sens2 and it's antisens peptides interactions are 1,143·10-6 M and 9,569·10-6
M. These results confirm sens and antisens peptides binding as well as potential antisens therapeutic effect which can
be further used in future studies. Also, positive results of this research suggest greater use of molecular recognition
theory for antisens peptide design which can be applied in various biomedicine fields.

• Keywords: Vipera ammodytes; poskok; amoditoksin A; teorija molekularnog prepoznavanja; sens; antisens; afinitet
• Keywords (english): Vipera ammodytes; viper; ammodytoxin A; molecular recognition theory; sens; antisens; affinity
• Language: croatian
• URN:NBN: urn:nbn:hr:163:951925
• Study programme: Title: Pharmacy; Study programme type: university; Study level: integrated undergraduate and graduate; Academic / professional title: magistar/magistra farmacije (magistar/magistra farmacije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2022-02-09 11:37:02