Abraham

Armougom

Bailey

Bell

Berendsen

Bird

Biswas

Boshoff

Brézellec

Collaborative Computational Project, Number 4

Corn

Darden

Davey

DeLano

Dhakaram Pangeni Sharma

Emsley

Fang

Grosset

Hacker

Hasegawa

Hess

Kashav

Katayama

Kornberg

Kumari

Laskowski

LeBowitz

Mann

Mitkova

Mott

Murshudov

O'Donnell

Oakley

Otwinowski

Panjikar

Parrinello

Perrakis

Ramachandran Vijayan

Rehman

Samudrala Gourinath

Schneider

Sheldrick

Soni

Syed Arif Abdul Rehman

Syson

Terwilliger

Tougu

Van Eijk

Velten

Wayne

Winn

Yamamoto

Zhang

The helicase-primase interaction is an essential event in DNA replication and is mediated by the highly variable C-terminal domain of primase (DnaG) and N-terminal domain of helicase (DnaB). To understand the functional conservation despite the low sequence homology of the DnaB-binding domains of DnaGs of eubacteria, we determined the crystal structure of the helicase-binding domain of DnaG from Mycobacterium tuberculosis (MtDnaG-CTD) and did so to a resolution of 1.58 Å. We observed the overall structure of MtDnaG-CTD to consist of two subdomains, the N-terminal globular region (GR) and the C-terminal helical hairpin region (HHR), connected by a small loop. Despite differences in some of its helices, the globular region was found to have broadly similar arrangements across the species, whereas the helical hairpins showed different orientations. To gain insights into the crucial helicase-primase interaction in M. tuberculosis, a complex was modeled using the MtDnaG-CTD and MtDnaB-NTD crystal structures. Two nonconserved hydrophobic residues (Ile605 and Phe615) of MtDnaG were identified as potential key residues interacting with MtDnaB. Biosensor-binding studies showed a significant decrease in the binding affinity of MtDnaB-NTD with the Ile605Ala mutant of MtDnaG-CTD compared with native MtDnaG-CTD. The loop, connecting the two helices of the HHR, was concluded to be largely responsible for the stability of the DnaB-DnaG complex. Also, MtDnaB-NTD showed micromolar affinity with DnaG-CTDs from Escherichia coli and Helicobacter pylori and unstable binding with DnaG-CTD from Vibrio cholerae The interacting domains of both DnaG and DnaB demonstrate the species-specific evolution of the replication initiation system.</p

Sharm, Dhakaram Pangeni

Vijayan, Ramachandran

Rehman, Syed Arif Abdul

Gourinath, Samudrala

Discovery Research Portal

Structural insights into the interaction of helicase and primase in Mycobacterium tuberculosis

English

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https://discovery.dundee.ac.uk/files/28723747/BCJ20180673.full.pdf

Structural insights into the interaction of helicase and primase in Mycobacterium tuberculosis

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