TY - JOUR

T1 - Conserved helicase domain of human RecQ4 is required for strand annealing-independent DNA unwinding

AU - Rossi, Marie L

AU - Ghosh, Avik K

AU - Kulikowicz, Tomasz

AU - Croteau, Deborah L

AU - Bohr, Vilhelm A

N1 - Published by Elsevier B.V.

PY - 2010/7/1

Y1 - 2010/7/1

N2 - Humans have five members of the well conserved RecQ helicase family: RecQ1, Bloom syndrome protein (BLM), Werner syndrome protein (WRN), RecQ4, and RecQ5, which are all known for their roles in maintaining genome stability. BLM, WRN, and RecQ4 are associated with premature aging and cancer predisposition. Of the three, RecQ4's biological and cellular roles have been least thoroughly characterized. Here we tested the helicase activity of purified human RecQ4 on various substrates. Consistent with recent results, we detected ATP-dependent RecQ4 unwinding of forked duplexes. However, our results provide the first evidence that human RecQ4's unwinding is independent of strand annealing, and that it does not require the presence of excess ssDNA. Moreover, we demonstrate that a point mutation of the conserved lysine in the Walker A motif abolished helicase activity, implying that not the N-terminal portion, but the helicase domain is solely responsible for the enzyme's unwinding activity. In addition, we demonstrate a novel stimulation of RecQ4's helicase activity by replication protein A, similar to that of RecQ1, BLM, WRN, and RecQ5. Together, these data indicate that specific biochemical activities and protein partners of RecQ4 are conserved with those of the other RecQ helicases.

AB - Humans have five members of the well conserved RecQ helicase family: RecQ1, Bloom syndrome protein (BLM), Werner syndrome protein (WRN), RecQ4, and RecQ5, which are all known for their roles in maintaining genome stability. BLM, WRN, and RecQ4 are associated with premature aging and cancer predisposition. Of the three, RecQ4's biological and cellular roles have been least thoroughly characterized. Here we tested the helicase activity of purified human RecQ4 on various substrates. Consistent with recent results, we detected ATP-dependent RecQ4 unwinding of forked duplexes. However, our results provide the first evidence that human RecQ4's unwinding is independent of strand annealing, and that it does not require the presence of excess ssDNA. Moreover, we demonstrate that a point mutation of the conserved lysine in the Walker A motif abolished helicase activity, implying that not the N-terminal portion, but the helicase domain is solely responsible for the enzyme's unwinding activity. In addition, we demonstrate a novel stimulation of RecQ4's helicase activity by replication protein A, similar to that of RecQ1, BLM, WRN, and RecQ5. Together, these data indicate that specific biochemical activities and protein partners of RecQ4 are conserved with those of the other RecQ helicases.

KW - Amino Acid Motifs

KW - Cell Nucleus

KW - Conserved Sequence

KW - DNA

KW - DNA Repair

KW - Humans

KW - Lysine

KW - Nucleic Acid Conformation

KW - Point Mutation

KW - Protein Structure, Tertiary

KW - RecQ Helicases

KW - Substrate Specificity

U2 - 10.1016/j.dnarep.2010.04.003

DO - 10.1016/j.dnarep.2010.04.003

M3 - Journal article

C2 - 20451470

VL - 9

SP - 796

EP - 804

JO - DNA Repair

JF - DNA Repair

SN - 1568-7864

IS - 7

ER -