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BioMed Research International
Volume 2014 (2014), Article ID 573936, 16 pages
http://dx.doi.org/10.1155/2014/573936
Research Article

C-Terminal Domain Swapping of SSB Changes the Size of the ssDNA Binding Site

1School of Biomedical Sciences, Chung Shan Medical University, No.110, Sec.1, Chien-Kuo N. Rd., Taichung City, Taiwan
2Department of Medical Research, Chung Shan Medical University Hospital, No.110, Sec.1, Chien-Kuo N. Rd., Taichung City, Taiwan

Received 26 May 2014; Accepted 9 July 2014; Published 4 August 2014

Academic Editor: Huangen Ding

Copyright © 2014 Yen-Hua Huang and Cheng-Yang Huang. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Single-stranded DNA-binding protein (SSB) plays an important role in DNA metabolism, including DNA replication, repair, and recombination, and is therefore essential for cell survival. Bacterial SSB consists of an N-terminal ssDNA-binding/oligomerization domain and a flexible C-terminal protein-protein interaction domain. We characterized the ssDNA-binding properties of Klebsiella pneumoniae SSB (KpSSB), Salmonella enterica Serovar Typhimurium LT2 SSB (StSSB), Pseudomonas aeruginosa PAO1 SSB (PaSSB), and two chimeric KpSSB proteins, namely, KpSSBnStSSBc and KpSSBnPaSSBc. The C-terminal domain of StSSB or PaSSB was exchanged with that of KpSSB through protein chimeragenesis. By using the electrophoretic mobility shift assay, we characterized the stoichiometry of KpSSB, StSSB, PaSSB, KpSSBnStSSBc, and KpSSBnPaSSBc, complexed with a series of ssDNA homopolymers. The binding site sizes were determined to be , , , , and nucleotides (nt), respectively. Comparison of the binding site sizes of KpSSB, KpSSBnStSSBc, and KpSSBnPaSSBc showed that the C-terminal domain swapping of SSB changes the size of the binding site. Our observations suggest that not only the conserved N-terminal domain but also the C-terminal domain of SSB is an important determinant for ssDNA binding.