Importance of the N-terminal Region of the Phage GA-1
Single-stranded DNA-binding Protein for Its Self-interaction Ability
and Functionality*
Irene
Gascón
,
José L.
Carrascosa§,
Laurentino
Villar,
José M.
Lázaro, and
Margarita
Salas¶
From the Centro de Biología Molecular
"Severo Ochoa" (Consejo Superior de Investigaciones
Científicas-Universidad Autónoma de Madrid) and
the § Centro Nacional de Biotecnología (Consejo
Superior de Investigaciones Científicas), Universidad
Autónoma, Cantoblanco, 28049 Madrid, Spain
The single-stranded DNA-binding protein (SSB) of
phage GA-1 displays higher efficiency than the SSBs of the related
phages 
29 and Nf. In this work, the self-interaction ability of GA-1 SSB has been analyzed by visualization of the purified protein by
electron microscopy, glycerol gradient sedimentation, and in vivo cross-linking of bacterial cultures infected with phage
GA-1. GA-1 SSB contains an insert at its N-terminal region that is not present in the SSBs of 29 and Nf. Three deletion mutant proteins have been characterized, 
N19, N26, and N33, which lack the 19, 26 or 33 amino acids, respectively, that follow the initial methionine
of GA-1 SSB. Mutant protein N19 retains the structural and
functional behavior of GA-1 SSB, whereas mutant proteins N26 and
N33 no longer stimulate viral DNA replication or display helix-destabilizing activity. Analysis of the mutant proteins by
ultracentrifugation in glycerol gradients and electron microscopy indicates that deletion of 26 or 33 but not of 19 amino acids of the
N-terminal region of GA-1 SSB results in the loss of the oligomerization ability of this protein. Our data support the importance of the N-terminal region of GA-1 SSB for the differential self-interaction ability and functional behavior of this protein.
*
This work was supported by Grant 2R01 GM27242-23 from the
National Institutes of Health, Grants PB-98-0645 and PB96-818 from Dirección General de Investigación Científica y
Técnica, and an institutional grant from Fundación
Ramón Areces to the Centro de Biología Molecular
"Severo Ochoa."The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
¶
To whom correspondence should be addressed. Tel.:
34-91-397-8435; Fax: 34-91-397-8490; E-mail:
msalas@cbm.uam.es.
Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.
