Conformational and Thermodynamic Consequences of the Introduction of a Nick in Duplexed DNA Fragments: an NMR Study Augmented by Biochemical Experiments

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 1989 Jun 26

PMID 2748334

Citations 16

Authors

J M Pieters

R M Mans

H van den Elst

G A van der Marel

J H VAN Boom

C Altona

Affiliations

Soon will be listed here.

Abstract

NMR studies were carried out on various equimolar mixtures consisting of a combination of oligomers: d(ACGGCT) (I). d(pACGGCT) (Ia), d(TGCAGT) (II), d(AGCCGTACTGCA) (III), d(TGCAGTACGGCT) (IV). It is shown that I + II + III (MI) and Ia + II + III (M2) form stable duplexes with nicks in the centre of the respective double helices. A close analysis of the NOESY experiments of M1 and M2 revealed that these fragments form B-DNA type duplex structures. A comparison of the chemical-shift data of the nicked duplexes with those of the intact duplex of III + IV (M3) demonstrated that only small local distortions occur when a nick is introduced. The chemical-shift profiles of M1 and M3 were used to obtain the thermodynamic data for the duplex/coil transitions. The profiles of M1 were analysed by means of a new thermodynamic model (TRIDUP). From the calculated thermodynamic data of M1 and M3 it is concluded that the melting behaviour of M1 occurs cooperatively. A ligation experiment demonstrated that the relatively small substrate (M2) was almost completely joined after an overnight incubation at 14 degrees C.

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