Plant Molecular Biology 33: 947–952, 1997.
1997 Kluwer Academic Publishers. Printed in Belgium.
Unraveling DNA helicases from plant cells
International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi 110 067, India
Received 31 December 1996; accepted in revised form 14 January 1997
Key words: DNA helicase, DNA-dependent ATPase, unwinding enzyme, DNA replication, chloroplasts
leus, mitochondria and chloroplasts, which have their
expresses its genetic information independent of each
other though there is (probably yet unknown) coordin-
ation between them. DNA present in these genomes
is in the double-stranded (ds) form. However, for
many important DNA transactions, such as replica-
tion, repair, recombination and insomecasestranscrip-
tion, the duplex DNA has to be partially unwind for
a short time to create single-stranded (ss) DNA inter-
mediates, through which all the proteins and enzymat-
ic machinery responsible for the above processes can
enter. A class of enzymes, called DNA helicases, cata-
lyzestheunwindingofduplexDNAin an ATPdepend-
ent fashion to provide the ssDNA template for such
processes and thus play an essential role in all aspects
of DNA metabolism.
The ﬁrst review of DNA helicases was published in
1981 by Geider and Hoffmann-Berlin . Since then
at least eight more reviews have been published [17–
20, 33, 34, 40, 44]. Plant DNA helicases have not
been covered in any of these reviews. Although much
has been learned from prokaryotic systems, much
still remains to be discovered about the mechanism
of unwinding and the exact role of many eukaryotic
DNA helicases, including those of plants. This mini-
review provides a brief overview of DNA helicases
and an account of progress-to-date in plant DNA hel-
icases. Theprimaryobjectiveinwritingthis articleis to
introduce the subject and simultaneously to stimulate
the plant biologists to work on this important enzyme,
This paper is dedicated to Prof. Arturo Falaschi and Prof.
Krishna K. Tewari.
since this ﬁeld is almost untouchedby plant biologists.
A biochemical technique to determine the unwinding
activity is described ﬁrst.
The helicase assay measures the displacement of
labelled DNA from a partial duplex molecule as
described [19, 42]. The biochemical reaction for DNA
helicase catalyzed unwinding is shown in Fig. 1A.
P-labelled partial duplex DNA substrate
is incubated with a DNA helicase enzyme in the pres-
ence of divalent cation (mainly Mg
) and nucleotide
triphosphate (mainly ATP or dATP). The DNA hel-
icase substrate as shown in Fig. 1A was prepared as
described earlier . This DNA helicase substrate
contains both 3
ssDNA ﬂanking regions (i.e.
a ‘forked’ structure). The optimum concentrations of
salt and ATP differ for different DNA hel-
icases. After the reaction, the products were separated
by native polyacrylamidegel electrophoresis followed
by autoradiography. The displaced fragment which is
radioactively labelled moves faster than the annealed
The polarity or direction of unwinding by helicase
is deﬁned with respect to the strand of DNA on which
the enzyme is bound. All DNA helicases move uni-
directionally, either 3
and prefer to
initiate unwinding of duplex DNA containing a ﬂank-
ing ssDNA region, with the exception of Escheri-
chia coli RecBCD enzyme which does not require a
strates . Some helicases show a strong preference
of unwinding duplex DNA containing a 3
ﬂanking region, whereas others show a preference for
GR: 201001885, PIPS Nr. 132240 BIO2KAP
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