Modular organization of a Cdc6-like protein from the crenarchaeon Sulfolobus solfataricus.
In the present paper, we report that a Cdc6 (cell-division control)-like factor from the hyperthermophilic crenarchaeon Sulfolobus solfataricus (referred to as SsoCdc6-2) has a modular organization of its biological functions. A reliable model of the SsoCdc6-2 three-dimensional structure was built up, based on the significant sequence identity with the Pyrobaculum aerophylum Cdc6 (PaeCdc6), whose crystallographic structure is known. This allowed us to design two truncated forms of SsoCdc6-2: the DeltaC (residues 1-297, molecular mass 35 kDa) and the DeltaN (residues 298-400, molecular mass 11 kDa) proteins. The DeltaC protein contains the nucleotide-binding Rossmann fold and the Sensor-2 motif (Domains I and II in the PaeCdc6 structure), and retains the ability to bind and hydrolyse ATP. On the other hand, the DeltaN protein contains the C-terminal WH (winged helix)-fold (Domain III), and is able to bind DNA molecules and to inhibit the DNA helicase activity of the SsoMCM (mini-chromosome maintenance) complex, although with lesser efficiency with respect to the full-sized SsoCdc6-2. These results provide direct biochemical evidence that the Cdc6 WH-domain is responsible for DNA-binding and inhibition of MCM DNA helicase activity.
De Felice
Mariarita
M
Istituto di Biochimica delle Proteine, Consiglio Nazionale delle Ricerche, Via P. Castellino 111, 80131-Napoli, Italy.
Esposito
Luca
L
Pucci
Biagio
B
De Falco
Mariarosaria
M
Manco
Giuseppe
G
Rossi
Mosè
M
Pisani
Francesca M
FM
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D017398:Alternative Splicing / Q000235:genetics; D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000096:biosynthesis / Q000235:genetics; D018797:Cell Cycle Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D019847:Chromosomes, Archaeal / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004265:DNA Helicases / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011485:Protein Binding / Q000187:drug effects; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D016944:Sulfolobus / Q000737:chemistry* / Q000201:enzymology
D019843:Archaeal Proteins; C066503:CDC6 protein, S cerevisiae; D018797:Cell Cycle Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D000255:Adenosine Triphosphate; D004247:DNA; D000251:Adenosine Triphosphatases; D004265:DNA Helicases