Molecular architecture of the recombinant human MCM2-7 helicase in complex with nucleotides and DNA
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Molecular architecture of the recombinant human MCM2-7 helicase in complex with nucleotides and DNA. / Boskovic, Jasminka; Bragado-Nilsson, Elisabeth; Saligram Prabhakar, Bhargav; Yefimenko, Igor; Martínez-Gago, Jaime; Muñoz, Sergio; Méndez, Juan; Montoya, Guillermo.
In: Cell Cycle, Vol. 15, No. 18, 16.09.2016, p. 2431-40.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - Molecular architecture of the recombinant human MCM2-7 helicase in complex with nucleotides and DNA
AU - Boskovic, Jasminka
AU - Bragado-Nilsson, Elisabeth
AU - Saligram Prabhakar, Bhargav
AU - Yefimenko, Igor
AU - Martínez-Gago, Jaime
AU - Muñoz, Sergio
AU - Méndez, Juan
AU - Montoya, Guillermo
PY - 2016/9/16
Y1 - 2016/9/16
N2 - DNA replication is a key biological process that involves different protein complexes whose assembly is rigorously regulated in a successive order. One of these complexes is a replicative hexameric helicase, the MCM complex, which is essential for the initiation and elongation phases of replication. After the assembly of a double heterohexameric MCM2-7 complex at replication origins in G1, the 2 heterohexamers separate from each other and associate with Cdc45 and GINS proteins in a CMG complex that is capable of unwinding dsDNA during S phase. Here, we have reconstituted and characterized the purified human MCM2-7 (hMCM2-7) hexameric complex by co-expression of its 6 different subunits in insect cells. The conformational variability of the complex has been analyzed by single particle electron microscopy in the presence of different nucleotide analogs and DNA. The interaction with nucleotide stabilizes the complex while DNA introduces conformational changes in the hexamer inducing a cylindrical shape. Our studies suggest that the assembly of GINS and Cdc45 to the hMCM2-7 hexamer would favor conformational changes on the hexamer bound to ssDNA shifting the cylindrical shape of the complex into a right-handed spiral conformation as observed in the CMG complex bound to DNA.
AB - DNA replication is a key biological process that involves different protein complexes whose assembly is rigorously regulated in a successive order. One of these complexes is a replicative hexameric helicase, the MCM complex, which is essential for the initiation and elongation phases of replication. After the assembly of a double heterohexameric MCM2-7 complex at replication origins in G1, the 2 heterohexamers separate from each other and associate with Cdc45 and GINS proteins in a CMG complex that is capable of unwinding dsDNA during S phase. Here, we have reconstituted and characterized the purified human MCM2-7 (hMCM2-7) hexameric complex by co-expression of its 6 different subunits in insect cells. The conformational variability of the complex has been analyzed by single particle electron microscopy in the presence of different nucleotide analogs and DNA. The interaction with nucleotide stabilizes the complex while DNA introduces conformational changes in the hexamer inducing a cylindrical shape. Our studies suggest that the assembly of GINS and Cdc45 to the hMCM2-7 hexamer would favor conformational changes on the hexamer bound to ssDNA shifting the cylindrical shape of the complex into a right-handed spiral conformation as observed in the CMG complex bound to DNA.
KW - Journal Article
U2 - 10.1080/15384101.2016.1191712
DO - 10.1080/15384101.2016.1191712
M3 - Journal article
C2 - 27249176
VL - 15
SP - 2431
EP - 2440
JO - Cell Cycle
JF - Cell Cycle
SN - 1538-4101
IS - 18
ER -
ID: 172395859