Polymerization Dynamics of the Prophage-Encoded Actin-Like Protein AlpC Is Influenced by the DNA-Binding Adapter AlpA.

Aaron J Forde,Catriona Donovan,Andreas Klingl,Nadine Albrecht,Marc Bramkamp

doi:10.3389/fmicb.2017.01429

Aaron J Forde, Catriona Donovan + Show 3 more

Open Access

https://doi.org/10.3389/fmicb.2017.01429

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Abstract

The Corynebacterium glutamicum ATCC 13032 prophage CGP3 encodes an actin-like protein, AlpC that was shown to be involved in viral DNA transport and efficient viral DNA replication. AlpC binds to an adapter, AlpA that in turn binds to specific DNA sequences, termed alpS sites. Thus, the AlpAC system is similar to the known plasmid segregation system ParMRS. So far it is unclear how the AlpACS system mediates DNA transport and, whether AlpA and AlpC functionally interact. We show here that AlpA modulates AlpC filamentation dynamics in a dual way. Unbound AlpA stimulates AlpC filament disassembly, while AlpA bound to alpS sites allows for AlpC filament formation. Based on these results we propose a simple search and capture model that explains DNA segregation by viral AlpACS DNA segregation system.

Highlights

Maintenance and stable inheritance of genetic material is a fundamental problem in all living cells
For characterization of the biochemical properties of the prophage encoded actin-like protein AlpC and its adapter protein AlpA the corresponding genes were cloned into expression vectors to allow for overexpression in E. coli as described in the Section “Materials and Methods.”
AlpA and AlpC were produced as fluorescent fusions proteins (GFP-AlpC and mCherry-AlpA) and with a histidine-tag to allow for metalaffinity chromatography

Summary

Introduction

Maintenance and stable inheritance of genetic material is a fundamental problem in all living cells. Type I (ParA-like) are defined by ParA proteins characterized by a deviant walker A motif, type II relies on actin-like proteins ALPs (ParM like) and type III are tubulin (TubZ) based (Gerdes et al, 2010; Salje et al, 2010). Common to these partitioning systems is that they function as simple tripartite structures. The loaded adapter in turn binds to a nucleotide hydrolyzing protein that either serves as a molecular gradient or polymer giving direction to the DNA segregation process (Gerdes et al, 2010; Salje et al, 2010; Schumacher, 2012)

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