|
Author
|
Topic: F1-ATPase and helicase evolved from common ancestor
|
Doubting Thomas
Member
Member # 1214
|
posted 30. March 2004 15:33
From Dr. Tony Crofts at the University of Illinois-Champaign-Urbana:
Evolution of the F1-ATPase
It has been suggested that the ATP synthase structure is too complicated to have evolved without Divine aid. Since the enzyme must, in this view, have been designed by a Creator, the complexity is taken as evidence for a creationist perspective of the origin of life. As a counter to this view, we might explore the possibility that the a and b subunits of the F1-ATPase shared a common ancestor with the hexameric helicases (1).
The hexameric helicases are ubiquitous proteins (related enzymes are found on both sides of bacterial-archaean divide) involved in unwinding double-stranded DNA and RNA. They bind to the single stranded form, and "walk" along it, forcing an unwinding at the bifurcation point of the double strand. Molecular weight studies of the active enzyme, and examination of the subunit content shows that the active forms are hexameric. Electron microscopic imaging of helicases from sources ranging from viruses to eukaryotes shows a symmetrical hexameric structure with a central hole. The single strand of DNA or RNA is threaded through the hole. Not all enzymes with helicase activity are hexameric; - the hexameric helicases are one class. In the hexameric class, the association formed is dependent on cofactors, - nucleotides and Mg2+, - and trimeric complexes can also be formed. However, the active enzyme is hexameric. Helicases are often found associated in multienzyme complexes, but the helicase isolated from these complexes retains activity and hexameric association under active conditions. The hexamers can be readily dissociated and reassociated. Functional studies using mixed hexamers, with one or more copies being inactive mutant forms, suggest that all six are required for function. If one of the 6 monomers is specifically inhibited, for example by covalent binding of 8-azido-ATP, function is lost.
The hexameric helicases are NTP hydrolases. The nucleotide triphosphate favored varies between types, and also with the substrate polynucleotide. We will assume that ATP is used in the following discussion. In the helicase reaction ATP hydrolysis accompanies movement along the DNA. Rapid kinetic studies show that ATP hydrolysis at three of the nucleotide binding sites is orders of magnitude slower that the overall rate of catalysis (kcat). Only three of the monomers appear to be involved in binding to the DNA, and in catalytic turnover in ATP hydrolysis. It is thought that the binding of alternate subunits in the hexameric ring distorts the interfaces between adjacent monomers so that alternating sites are in a non-catalytic configuration. Of the three rapid sites, 1 ATP/hexamer is hydrolyzed faster than kcat, suggesting that the three are not equivalent. Since ssDNA is itself asymmetric in cross-section, the three subunits involved in binding will see a different face of the threaded strand. The picture that emerges from kinetic studies is of a mechanism in which the three catalytic subunits are involved in a sequential mechanism in which they change configuration to match different binding faces. Local changes in affinity for a periodic pattern along the DNA, with transient unbinding and rebinding of each subunit, would “force” a progress along the DNA. These reactions are mechanistically linked to ATP hydrolysis, which provides the driving force.
The parallel between the hexameric helicases and the F1 ATPase is quite striking. Both consist of hexameric rings, but the F1 ring is composed of 3 each of alternating a and b subunits. However, the sequence similarity and homologous tertiary structure strongly suggest that these two types evolved from a common ancestor, indicating that the ancestral structure was a homohexameric structure. The a and b subunits also show sequence and structural similarity to the hexameric helicase monomers, suggesting that they all share a common ancestry.
The a and b subunits form a ring around a centrally located g subunit, which interacts asymmetrically with the three b subunits. In the Walker structure, this asymmetry of interaction is associated with an asymmetry of occupation of the three catalytic sites on the b subunits. One contains the ATP analogue AMP-PNP, one ADP, and one is empty. The a subunits all contain nucleotide, - the ATP analogue AMP-PNP. The binding sites are more or less symmetrically located at the interfaces between a and b subunits, with “regulatory” a subunit and catalytic b subunit sites alternating around the hexameric ring. The complete structures show distinct binding sites along the g subunit, which interact differentially with the interfacial contact spans of each b subunit. Since the contact sites are on three faces reflecting the pseudo-C3 symmetry, the sequential changes in configuration allow the hexameric ring to rotate the g subunit by “walking” along it!
From the above description, it will be apparent that the F1 ATPase mechanism is essentially the same as the helicase mechanism. In view of the sequence and structural similarity, it seems quite likely therefore that the a, b-ring evolved from an ancestral homohexameric structure shared in common with the helicases. If the a, b-ring evolved, then the enzyme evolved.
It would be going beyond the evidence to claim that we understand the evolution of the ATP synthase. It must have happened before the archaeal and bacterial lines separated, in a process for which we have only the modern forms, and the tools of genomics to provide a guide. We can however claim that the complexity of the modern forms is not an indication that they were designed, or created in their present forms. They evolved from simpler precursors as would be expected from an evolutionary origin.
1. S. S. Patel and K. M. Picha (2000) Structure and function of hexameric helicases. Annu. Rev. Biochem. 69, 651-697
[ 31. March 2004, 20:05: Message edited by: Doubting Thomas ]
IP: Logged
|
|
Doubting Thomas
Member
Member # 1214
|
posted 31. March 2004 11:15
Nature 409, 573 - 575 (2001)
Structural biology: Pumping DNA
EDWARD H. EGELMAN1
Edward H. Egelman is in the Department of Biochemistry and Molecular Genetics, University of Virginia Health Sciences Center, Charlottesville, Virginia 22908, USA.
e-mail: egelman@virginia.edu
Crystal structures of proteins not only shed light on how those proteins work. By revealing previously hidden similarities, they can also force a re-evaluation of what other proteins are predicted to do.
Atomic structures of proteins often reveal evolutionary relationships between them that cannot be seen when looking at sequences of amino acids. A particular fold or topology might be preserved in two proteins from different species, or in different proteins from the same species, even though their amino-acid sequences have diverged so far during evolution that they have no recognizable similarities. This conservation of structure reinforces the idea that there is only a limited number of stable shapes that proteins can have. It also shows how studies of bacteria or viruses can provide an insight into some processes in higher organisms.
On page 637 of this issue1, Gomis-Rüth and colleagues show that a protein responsible for transferring single-stranded DNA between bacteria is part of a huge family of proteins involved in the replication, repair, recombination and expression of DNA, as well as in transforming energy. The discovery provides insight into how this protein might function as a DNA pump. It also tells us more about this ubiquitous protein superfamily.
The first member of the superfamily to have its structure solved was the protein RecA from the bacterium Escherichia coli2. This intensively studied enzyme is involved in homologous genetic recombination — the process by which two DNA molecules with similar sequences swap strands of DNA. Recombination is a way of repairing DNA, maintaining the stability of the genome, and generating genetic diversity. (In higher organisms, recombination occurs during the production of reproductive cells.) The main active form of RecA appears to be a helical filament formed on DNA, but the protein also forms rings containing six RecA proteins3.
The next protein to be included in this superfamily was a helicase: despite having very different amino-acid sequences, RecA and this helicase4 have the same core. Helicases use energy derived from the hydrolysis of ATP to separate double-stranded DNA or RNA into two single strands. These proteins were originally studied in the context of DNA replication — double-stranded DNA must be separated into two strands to allow access by the replicating machinery. But it is now known that helicases participate in all aspects of DNA metabolism, including recombination, replication, repair and transcription. An analysis of the genome of the yeast Saccharomyces cerevisiae showed that about 2% of its genes (at least 134 genes) encode helicases, as judged by the presence of certain sequence motifs5.
Subsequent structural studies have provided support for the prediction4 that all helicases contain the same core as RecA. And just as RecA can exist in a ring form, so too can some of the most well characterized helicases6. There is a growing consensus that many of these function by allowing one strand of DNA to pass through the centre of the ring while the complementary strand is displaced outside the ring7. However, a bacterial helicase called RuvB appears to function in recombination by pumping double-stranded DNA through the central channel of its hexameric ring8.
Another addition to this protein superfamily came as a greater surprise. The core of RecA and many helicases is also seen in the F1-ATPase9. This protein, found only in higher organisms, is part of the F1F0-ATP synthase, which makes ATP — the main source of energy for most cellular processes. So it seems that although the functions (and sequences) of proteins such as helicases and the F1-ATPase have diverged greatly, aspects of their organization into hexameric rings have been preserved.
Gomis-Rüth et al.1 now add the E. coli protein TrwB to the family. They have determined the structure of TrwB, and find that it exists as a hexameric ring that is remarkably similar to those formed by the F1-ATPase, RecA and certain helicases (Fig. 1). The structure and topology of the core regions are also very similar.
References 1. Gomis-Rüth, F. X. et al. Nature 409, 637-641 (2001). | Article | PubMed | ISI | ChemPort |
2. Story, R. M., Weber, I. T. & Steitz, T. A. Nature 355, 318-325 (1992). | Article | PubMed | ISI | ChemPort |
3. Yu, X. & Egelman, E. H. Nature Struct. Biol. 4, 101-104 (1997). | PubMed | ISI | ChemPort |
4. Subramanya, H. S. et al. Nature 384, 379-383 (1996). | Article | PubMed | ISI | ChemPort |
5. Shiratori, A. et al. Yeast 15, 219-253 (1999). | Article | PubMed | ISI | ChemPort |
6. Singleton, M. R. et al. Cell 101, 589-600 (2000). | PubMed | ISI | ChemPort |
7. Yu, X. et al. Nature Struct. Biol. 3, 740-743 (1996). | PubMed | ISI | ChemPort |
8. Parsons, C. A. et al. Nature 374, 375-378 (1995). | Article | PubMed | ISI | ChemPort |
9. Abrahams, J. P. et al. Nature 370, 621-628 (1994). | Article | PubMed | ISI | ChemPort |
10. Bath, J. et al. Science 290, 995-997 (2000). | Article | PubMed | ISI | ChemPort
IP: Logged
|
|
|
Printer-friendly view of this topic