The Sulfolobus solfataricus GINS Complex Stimulates DNA Binding and Processive DNA Unwinding by Minichromosome Maintenance Helicase.
GINS is a key component of the eukaryotic Cdc45-minichromosome maintenance (MCM)-GINS (CMG) complex, which unwinds duplex DNA at the moving replication fork. Archaeal GINS complexes have been shown to stimulate the helicase activity of their cognate MCM mainly by elevating its ATPase activity. Here, we report that GINS from the thermoacidophilic crenarchaeon Sulfolobus solfataricus (SsoGINS) is capable of DNA binding and binds preferentially to single-stranded DNA (ssDNA) over double-stranded DNA (dsDNA). Notably, SsoGINS binds more strongly to dsDNA with a 5' ssDNA tail than to dsDNA with a 3' tail and more strongly to an ssDNA fragment blocked at the 3' end than to one at the 5' end with a biotin-streptavidin (SA) complex, suggesting the ability of the protein complex to slide in a 5'-to-3' direction along ssDNA. DNA-bound SsoGINS enhances DNA binding by SsoMCM. Furthermore, SsoGINS increases the helicase activity of SsoMCM. However, the ATPase activity of SsoMCM is not affected by SsoGINS. Our results suggest that SsoGINS facilitates processive DNA unwinding by SsoMCM by enhancing the binding of the helicase to DNA. We propose that SsoGINS stabilizes the interaction of SsoMCM with the replication fork and moves along with the helicase as the fork progresses.


OBJECTIVE
GINS is a key component of the eukaryotic Cdc45-MCM-GINS complex, a molecular motor that drives the unwinding of DNA in front of the replication fork. Archaea also encode GINS, which interacts with MCM, the helicase. But how archaeal GINS serves its role remains to be understood. In this study, we show that GINS from the hyperthermophilic archaeon Sulfolobus solfataricus is able to bind to DNA and slide along ssDNA in a 5'-to-3' direction. Furthermore, Sulfolobus GINS enhances DNA binding by MCM, which slides along ssDNA in a 3'-to-5' direction. Taken together, these results suggest that Sulfolobus GINS may stabilize the interaction of MCM with the moving replication fork, facilitating processive DNA unwinding.
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.
Shiwei
S
Lang
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China huangl@sun.im.ac.cn.
Li
L
Huang
D019843:Archaeal Proteins / Q000378:metabolism; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication; D019641:DNA, Archaeal / Q000235:genetics* / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D064110:Minichromosome Maintenance Proteins / Q000235:genetics / Q000378:metabolism*; D048229:Sulfolobus solfataricus / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism
D019843:Archaeal Proteins; D019641:DNA, Archaeal; D004277:DNA, Single-Stranded; D004265:DNA Helicases; D064110:Minichromosome Maintenance Proteins