Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of the central zinc-binding domain of the human Mcm10 DNA-replication factor.
The initiation of eukaryotic DNA replication requires the tightly controlled assembly of a set of replication factors. Mcm10 is a highly conserved nuclear protein that plays a key role in the initiation and elongation processes of DNA replication by providing a physical link between the Mcm2-7 complex and DNA polymerases. The central domain, which contains the CCCH zinc-binding motif, is most conserved within Mcm10 and binds to DNA and several proteins, including proliferative cell nuclear antigen. In this study, the central domain of human Mcm10 was crystallized using the hanging-drop vapour-diffusion method in the presence of PEG 3350. An X-ray diffraction data set was collected to a resolution of 2.6 A on a synchrotron beamline. The crystals formed belonged to space group R3, with unit-cell parameters a = b = 99.5, c = 133.0 A. According to Matthews coefficient calculations, the crystals were predicted to contain six MCM10 central domain molecules in the asymmetric unit.
National Creative Initiatives for Structural Biology and Department of Life Science, Pohang University of Science and Technology, Hyo-ja dong, San31, Pohang, KyungBook, Republic of Korea.
Nam Young
NY
Jung
Won Jin
WJ
Bae
Jeong Ho
JH
Chang
Young Chang
YC
Kim
Yunje
Y
Cho
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000032:analysis* / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D002460:Cell Line; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D005822:Genetic Vectors; D006801:Humans; D007668:Kidney / Q000166:cytology; D064110:Minichromosome Maintenance Proteins; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques; D017434:Protein Structure, Tertiary; D014961:X-Ray Diffraction
D018797:Cell Cycle Proteins; D018076:DNA, Complementary; C489613:MCM10 protein, human; D064110:Minichromosome Maintenance Proteins