Abstract
Author SummaryLinear chromosomes capped by hairpin telomeres are widespread in prokaryotes and are found in important bacterial pathogens. However, three-dimensional structure of the hairpin telomere, as well as the molecular mechanisms underlying its generation, has remained poorly understood. In this work, we investigated how the enzyme responsible for generating the bacterial hairpin telomeres (protelomerase, also known as telomere resolvase) transforms a linear double-stranded DNA molecule into sharp hairpin turns. Our X-ray crystallographic and biochemical data collectively suggest that protelomerase employs a multistep DNA strand-refolding mechanism as described below. Protelomerase first cleaves both strands of a double-helical DNA substrate and reshapes the DNA strands into a transition state conformation (refolding intermediate) stabilized by specific protein–DNA and DNA–DNA interactions including noncanonical (non-Watson–Crick) base-pairs. The DNA strands are then refolded into extremely compact hairpin products, stabilized by a set of interactions distinct from those stabilizing the refolding intermediate. We believe that an enzyme “catalyzing” not only the chemical reactions of DNA strand cutting/rejoining but also the ordered transition between different DNA conformations to guide refolding of the DNA strand is a novel concept, and we suspect that similar mechanisms may be employed by other enzymes involved in conformational changes/refolding of biological macromolecules.
Highlights
Telomeres at the termini of linear chromosomes protect the deoxyribonucleic acid (DNA) ends from degradation or aberrant repair reactions including end-fusion, while allowing complete replication of the terminal sequences [1]
Overall Structure of the TelA–DNA Complex To better understand how protelomerase functions, we have crystallized the full-length protelomerase from the plant pathogen Agrobacterium tumefaciens C58 (TelA) in complex with DNA substrates containing the terminal sequences of the Agrobacterium linear chromosome
Phases for a parental TelA–DNA complex were obtained by the selenomethionine single-wavelength anomalous dispersion (SAD) phasing method, and all crystal structures have been refined to 2.2,2.4 Aresolution
Summary
Telomeres at the termini of linear chromosomes protect the DNA ends from degradation or aberrant repair reactions including end-fusion, while allowing complete replication of the terminal sequences [1]. The simplest form of telomere is a covalently closed hairpin loop found in bacteria carrying linear chromosomes, including Borrelia spirochetes—the causative agents of Lyme disease and relapsing fever [2,3], Agrobacterium [4,5], and cyanobacteria. Replication of such bacterial linear chromosomes with hairpin telomeres starts from an internal origin and proceeds bi-directionally, yielding a circular intermediate composed of a head-to-head, tail-to-tail dimer of chromosomes (Figure 1A) [6,7]. Mechanisms by which protelomerase drives the hairpin formation reaction forward without an exogenous input of energy or getting trapped in a DNA cleavage–religation equilibrium are not well understood
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