Citation:

Optical and Structural Characterization of a Chronic Myeloid Leukemia DNA Biosensor, Cordeiro, Mílton, Otrelo-Cardoso {Ana Rita Castro}, Svergun {Dmitri I. }, Konarev {Petr V. }, Lima {João Carlos}, Santos-Silva Teresa, and Baptista {Pedro Viana} , ACS Chemical Biology, may, Volume 13, Number 5, p.1235–1242, (2018)

Abstract:

Selective base pairing is the foundation of DNA recognition. Here, we elucidate the molecular and structural details of a FRET-based two-component molecular beacon relying on steady-state fluorescence spectroscopy, small-angle X-ray scattering (SAXS), microscale thermophoresis (MST), and differential electrophoretic mobility. This molecular beacon was designed to detect the most common fusion sequences causing chronic myeloid leukemia, e14a2 and e13a2. The emission spectra indicate that the self-assembly of the different components of the biosensor occurs sequentially, triggered by the fully complementary target. We further assessed the structural alterations leading to the specific fluorescence FRET signature by SAXS, MST, and the differential electrophoretic mobility, where the size range observed is consistent with hybridization and formation of a 1:1:1 complex for the probe in the presence of the complementary target and revelator. These results highlight the importance of different techniques to explore conformational DNA changes in solution and its potential to design and characterize molecular biosensors for genetic disease diagnosis.

Notes:

Sem PDF conforme despacho. info:eu-repo/grantAgreement/FCT/5876/147258/PT# info:eu-repo/grantAgreement/FCT/SFRH/SFRH%2FBD%2F85806%2F2012/PT# This work was supported by the Unidade de Cienciaŝ Biomoleculares Aplicadas-UCIBIO, which is financed by national funds from FCT/MEC (UID/Multi/04378/2013) and cofinanced by the ERDF under the PT2020 Partnership Agreement (POCI-01-0145-FEDER-007728) and PTDC/ QEQ-MED/1902/2014. A.R.O.-C. and M.C. thank FCT/ MEC for SFRH/BD/85806/2012 and SFRH/BD/87836/ 2012, respectively. The authors thank the EMBL P12 (DESY, Hamburg, Germany) and BM29 (ESRF, Grenoble, France) beamline staff for assistance during data collection. P.V.K. acknowledges the support from Federal Agency of Scientific Organizations (Agreement No 007-Γ3/43363/26).; 6th Annual Conference of the International Chemical Biology Society (ICBS), ICBS ; Conference date: 17-10-2017 Through 20-10-2017