Helicases as molecular motors: An insight

Abstract

Helicases are one of the smallest motors of biological system, which harness the chemical free energy of ATP hydrolysis to catalyze the opening of energetically stable duplex nucleic acids and thereby are involved in almost all aspect of nucleic acid metabolism including replication, repair, recombination, transcription, translation, and ribosome biogenesis. Basically, they break the hydrogen bonding between the duplex helix and translocate unidirectionally along the bound strand. Mostly all the helicases contain some conserved signature motifs, which act as an engine to power the unwinding. After the discovery of the first prokaryotic DNA helicase from Escherichia coli bacteria in 1976 and the first eukaryotic one from the lily plant in 1978, many more (>100) have been isolated. All the helicases share some common properties, including nucleic acid binding, NTP hydrolysis and unwinding of the duplex. Many helicases have been crystallized and their structures have revealed an underlying common structural fold for their function. The defects in helicases gene have also been reported to be responsible for variety of human genetic disorders, which can lead to cancer, premature aging or mental retardation. Recently, a new role of a helicase in abiotic stress signaling in plant has been discovered. Overall, helicases act as essential molecular tools for cellular machinery and help in maintaining the integrity of genome. Here an overview of helicases has been covered which includes history, biochemical assay, properties, classification, role in human disease and mechanism of unwinding and translocation.

Keywords: ATPase; Helicase classification; Helicase-related diseases; Molecular motors; Signature motifs; Unwinding enzyme.

Fig. 1

A. Scheme of biochemical assay for measuring unwinding activity of ATP/Mg²⁺-dependent DNA helicase. Asterisks denote the ³²P-labeled end of the DNA. The partial duplex DNA helicase substrate was prepared by annealing the radiolabeled DNA oligo (small DNA) to longer ssDNA as described , . B. Autoradiogram of the gel: lane 1, reaction without enzyme; lane 2, heat-denatured substrate and lane 3, reaction in presence of DNA helicase enzyme. S=substrate; UD=unwound DNA.

Fig. 2

Models for mechanism of DNA helicase unwinding and translocation. The helicase protein contacts and translocates on the sugar-phosphate backbone of the DNA strand and the hydrolysis of ATP is required . A. Active rolling model: The two subunits of dimeric helicase are shown as oval shape in which one is black. The dimeric helicase unwinds by interacting directly with both duplex and ssDNA. Each subunit alternates binding to duplex DNA as the dimer translocates when one subunit releases ssDNA and rebinds to duplex DNA . In this model translocation along ssDNA is coupled to ATP binding, whereas ATP hydrolysis drives the unwinding of multiple DNA base pairs for each catalytic event. B. Inchworm model: This model is consistent with monomeric or oligomeric state for the protein (shown as oval shape). The enzyme monomer first binds to ssDNA and then translocates along the DNA strand and then binds to the duplex region at fork followed by unwinding and release of one of the ssDNA strand . If the enzyme is hexamer as SV40 large T antigen, then one of the subunit (monomer 1) remains associated with the fork through the unwinding cycle . C. Modified inchwarm model: This model is proposed for monomeric UvrD helicase . In this model, it is assumed that the monomeric helicase contain two DNA binding sites: the leading site (L) binds to ds- and ssDNA both and the trailing site (T) binds to only ssDNA. Upon ATP binding the enzyme changes its conformation from extended to compact state, in which the T site is shifted forward along the ssDNA towards the L site. Upon ATP hydrolysis the enzyme changes its conformation from compact to extended state. In the extended state the T site is bound to ssDNA while L site is extended forward in the duplex region and unwinds the DNA. In all the models the shape of the helicase and location of the DNA are arbitrary .