Effects of single amino acid substitutions at the predicted coiled-coil or hydrophobic region on the self-assembly of ϕ29 replication protein, gp1

Single-Stranded DNA Binding Protein Gp5 ofBacillus subtilisPhage Φ29 Is Required for Viral DNA Replication in Growth-Temperature Dependent Fashion

“…When gp5-producing B. subtilis TT299 was infected with ns5, an increase in viral DNA was observed. These results indicate that in the absence of functional gp5, viral DNA replication is blocked at 45 C. To date, two '29 temperature-sensitive mutants in gene 5, ts5 (17) and ts5(219), have been isolated. 3,24) Using ts5 (17), gp5 has been found to be essential for viral DNA replication in vivo.…”

“…These results indicate that in the absence of functional gp5, viral DNA replication is blocked at 45 C. To date, two '29 temperature-sensitive mutants in gene 5, ts5 (17) and ts5(219), have been isolated. 3,24) Using ts5 (17), gp5 has been found to be essential for viral DNA replication in vivo. [3][4][5] However, the sequence of the gene 5 mutation was not reported, and we did not find any mutation in the gene 5 in our laboratory stock of ts5 (17).…”

“…We have studied the function of gene 1, [17][18][19][20][21] a viral gene involved in '29 DNA replication. In a genetic study using gene 1 mutants, we identified several gene 5 mutations that suppress temperature sensitive growth caused by gene 1 mutations (manuscript in preparation).…”

Isolation of suppressors of the temperature-sensitive growth caused by a nonsense mutation in gene 1 of <i>Bacillus subtilis</i> phage ø29 using hydroxylamine

“…2B). The C-terminal region of gp1 is composed of the putative coiled-coil and the hydrophobic regions which were reported to be important for the self-association of gp1 (Bravo and Salas, 1998;Hashiyama et al, 2005). Furthermore, no ortholog of ø29 gene 1 has been found in the genome of GA-1 (Meijer et al, 2001).…”

A Variant Protein from φ29 Replication Gene, Gene 1, Did Not Form Homo-Polymer Due to a Single Amino Acid Substitution Near the Carboxyl Terminus

“…While its self-association was suppressed, the product of the W71R type gene 1 was found to retain RNA binding activity. 5,11) Therefore, the variants with amino acid substitutions within the hydrophobic region ought to be powerful for detailed investigation of the function of gp1 and for the analysis of its three-dimensional structure. Purified and refolded gp1s from wild-type or W71R gene 1 were incubated with gp1 without a His-tag (from the wild-type or the W71R type gene 1) for 2 or 5 h at 4 C in 50 ml of the refolding buffer without urea (Fig.…”