SPMS MS02 02 Sin ingle le-Mo Molec lecule ule Bi Biophys - - PowerPoint PPT Presentation

SPMS MS02 02

Sin ingle le-Mo Molec lecule ule Bi Biophys hysics: ics: Mec Mecha hani nica cal l Un Unfo fold ldin ing g of R f RNA NA by y Optica ical l Tw Twee eezer ers

In Introduc

• ducti

tion

• n

Objective:

Investigate a potential RNA triplex structure stimulating ribosomal frameshifting Mechanical pulling of target RNA in different chemical environments using optical tweezer

through

enables recoding of ribosomal frameshifting

important structural motif found in pseudoknots and structured RNA

Tr Trip iplex lex RNA NA

serves catalytic and regulatory functions in cellular nucleoprotein complexes forms through tertiary interactions in the major or minor groove of a Watson-Crick base-paired stem

Rib ibos

• somal
• mal Fr

Fram ameshift eshifting ing

Op Opti tical al Tw Twee eezer er

Bu Buffers ffers Us Used ed

(i) 200 mM NaCl, 20 mM NaOAc, 0.1 mM EDTA, pH 5.0 and (ii) 5 mM MgCl2, 100 μM spermine, 200 mM NaCl, 20 mM NaOAc, 0.1 mM EDTA, pH 5.0.

lowered pH is in favour of the formation

• f the triplex structure

Pu Pull llin ing g Mod

• des

es

Pulling Modes

Force-ramp Constant-force

Pu Pull llin ing g Mod

• de:

e: Fo Force-ramp amp

Pu Pull llin ing g Mod

• de:

e: Co Constan tant-force force

Diagram of raw data for forces 16.5pN, 16.8pN, 17.1pN and 17.4pN in buffer 1 Diagram of raw data for forces 17.4pN, 17.7pN, 18.0pN and 18.3pN in buffer 2

Buffer 1: Critical force= 17.0pN Buffer 2: Critical force= 18.1pN

Cr Crit itic ical al Fo Force

What at I h I hav ave e le lear arnt

The role of triplex in stabilising RNA The correlation between mechanical property and stability of RNA triplex Spermine and Mg2+ stabilize the RNA structure.

Im Impr provement

• vement

Melting studies done by Ultraviolet absorption Thermodynamic information (stability) on triplex RNA Melting temperature (Tm)

Th Than ank k yo you! u!