Functional Analysis of the Superfamily 1 DNA Helicases Encoded by Mycoplasma pneumoniae and Mycoplasma genitalium

Silvia Estevão,Cornelis Vink,Theo Hoogenboezem,Annemarie M C Van Rossum,Helga U Van Der Heul,Marcel Sluijter,Nico G Hartwig

doi:10.1371/journal.pone.0070870

Silvia Estevão, Cornelis Vink + Show 5 more

Open Access

https://doi.org/10.1371/journal.pone.0070870

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Journal: PloS one	Publication Date: Jul 23, 2013
Citations: 2	License type: CC BY 4.0

Affiliation: Erasmus MC - Sophia Children’s Hospital

Abstract

The DNA recombination and repair machinery of Mycoplasma pneumoniae is composed of a limited set of approximately 11 proteins. Two of these proteins were predicted to be encoded by neighboring open reading frames (ORFs) MPN340 and MPN341. Both ORFs were found to have sequence similarity with genes that encode proteins belonging to the DNA helicase superfamily 1 (SF1). Interestingly, while a homolog of the MPN341 ORF is present in the genome of Mycoplasma genitalium (ORF MG244), MPN340 is an M. pneumoniae-specific ORF that is not found in other mycoplasmas. Moreover, the length of MPN340 (1590 base pairs [bp]) is considerably shorter than that of MPN341 (2148 bp). Examination of the MPN340-encoded amino acid sequence indicated that it may lack a so-called 2B subdomain, which is found in most SF1 DNA helicases. Also, the MPN340-encoded amino acid sequence was found to differ between subtype 1 strain M129 and subtype 2 strain FH at three amino acid positions. Both protein variants, which were termed PcrAs M129 and PcrAs FH, respectively, as well as the MPN341- and MG244-encoded proteins (PcrAMpn and PcrAMge, respectively), were purified, and tested for their ability to interact with DNA. While PcrAMpn and PcrAMge were found to bind preferentially to single-stranded DNA, both PcrAs M129 and PcrAs FH did not demonstrate significant DNA binding. However, all four proteins were found to have divalent cation- and ATP-dependent DNA helicase activity. The proteins displayed highest activity on partially double-stranded DNA substrates carrying 3′ single-stranded extensions.

Highlights

Mycoplasma pneumoniae and Mycoplasma genitalium are genetically closely related human pathogens that are classified within the bacterial class of Mollicutes
To increase the understanding of the functionality of the ‘minimal’ DNA recombination and repair (DRR) machinery of the pathogenic mycoplasmas, we have focused our attention on the M. pneumoniae and M. genitalium open reading frames (ORFs) that share sequences with genes encoding PcrA/Rep/ UvrD-like DNA helicases
The alignment demonstrated that the MPN340, MPNE_0394, MPN341- and MG244-encoded sequences each have features characteristic of proteins belonging to the SF1A group from the superfamily 1 (SF1) superfamily of DNA helicases [22,23,24,34]

Summary

Introduction

From in silico analyses of mycoplasma genomes it was predicted that they possess the most compact set of recombination-associated genes of all known bacteria, consisting of approximately 11 genes [5,6] These genes have the capacity to code for proteins putatively involved in homologous DNA strand transfer (RecA and SSB), Holliday junction (HJ) branch migration and resolution (RuvA, RuvB and RecU), nucleotide excision repair (UvrA, UvrB, UvrC, and PcrA) and base excision repair (Nfo endonuclease IV). In comparison to other bacteria, M. pneumoniae and M. genitalium appear to lack homologs of several other enzymes and/or enzymatic pathways involved in DRR Enzymes such as LexA and others related to the SOS response are absent. Despite the apparent limitations of their DRR machineries, homologous DNA recombination events were found to occur in both M. pneumoniae and M. genitalium [7,8,9,10,11]

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