Reactive walls

Abstract

Domain walls are natural borders in ferromagnetic, ferroelectric or ferroelastic materials. It seems that they can also be reactive areas that produce crystallographic phases never before observed in bulk materials. See Letter p.379 polymerization, involves the chemical joining of ribonucleotide monomers or oligonucleo-tides (short sequences of monomers). Some 30 years ago, a conundrum arose concerning how RNA molecules first proliferated through prebiotic chemical reactions. This was because of the demonstration by Joyce et al. 3 that the non-enzymatic copying of an RNA template to form a complementary RNA strand could be brought to a screeching halt by the incorporation into the growing polymer of monomers of opposite handed-ness to the template. This phenomenon was termed 'enantio meric cross-inhibition'. Given that both d-and l-enantiomers of RNA molecules were probably present as substrates on prebiotic Earth, how could template-directed polymerization have proceeded? Sczepanski and Joyce now revisit this issue by creating a ribozyme that not only catalyses template-directed polymerization in the presence of both d-and l-enantiomers, but actually prefers mononucleotides and oligonucleotides of the opposite handedness to itself as its substrates. The authors synthesized a pool of right-handed d-RNA polymers of random sequences and linked them covalently to a left-handed l-RNA template in the presence of left-handed oligonucleotide substrates. They then used in vitro selection 4-6 to isolate RNA species from the pool that could join (poly merize) the substrates. After ten rounds of selection and amplification of catalytic molecules; pruning of superfluous sequences; insertion of another randomized segment to create a new pool; and then another six rounds of selection and amplification, a d-ribozyme was isolated that could perform template-directed joining of l-substrates about a million times faster than in the uncatalysed reaction 1. As with ribozymes previously selected and further optimized for polymerization activity 7-9 , this ribozyme resembles modern-day polymeri-zation enzymes (polymerases) in several ways. First, it can operate on completely separate template-substrate complexes, implying that sequence-independent contacts form between it and the complexes. Second, it can perform limited polymerization by catalysing the sequential joining of several mononucleotides. In addition, as long as the oligomeric substrates are bound to their complementary templates, the ribozyme seems to be indifferent to sub-strate length. In fact, the authors observed that it can connect 11 l-oligo nucleotides to form a mirror copy of itself, a remarkable first demonstration of an enzyme (RNA or protein) being synthesized by its own enantiomer. Importantly, the d-ribozyme and its l-enantiomer efficiently catalyse their respective joining reactions even in a mixture containing both d-and l-versions of the substrates and templates. In other words, the enantiomeric cross-inhibition that thwarted non-enzymatic template-mediated replication does not occur. This work adds weight to the notion of a primordial RNA world in which cycles of cross-handed replication used mirror-image forms of RNA (Fig. 1). Such mutualistic coupling of d-and l-RNA polymerases might have conferred several advantages on RNA evolution, and may now benefit researchers who aspire to create RNA polymerase systems that can self-replicate. Because of their shapes, d-and l-RNA molecules cannot form consecutive Watson-Crick base pairs with each other 10 , just as left and right hands cannot properly handshake one another. Consequently, Sczepanski and Joyce's polymer-ase is unlikely to exhibit sequence preferences or restrictions owing to duplex formation between complementary sequences in the ribozyme and in templates or reaction products. These problems have confounded the experimental search for a general polymerase that can copy RNA sequences without bias, and may similarly have affected the course of early macromolecular evolution. Furthermore , the dependence on two distinct, coupled polymerases makes the replication cycle less susceptible to invasion by molecular parasites that could usurp the chemically activated sub-strates needed for the polymerization reaction. Viewing the work in the context of evolution begs the question of how an all d-or all l-ribozyme could have arisen to begin with. Sczepanski and Joyce suggest that simple forms of nucleic acids that lacked the mirror twin served as templates for polymerization of RNA mono-or oligonucleotide substrates on prebiotic Earth. It remains unclear, however, whether RNA polymerization from such templates would be immune to deleterious enan-tiomeric cross-inhibition. Beyond the implications for the RNA world hypothesis, the new ribozyme may have practical value for the production of spiegel-mers 11-l-versions of functional d-RNAs (the name derives from the German word for 'mirror': Spiegel). Spiegelmers resist degradation by nucleases-the enzymes that degrade nucleic acids-and seem to avoid detection by the immune system, making them attractive therapeutic candidates and sensors for biological ligands. However, because natural enzymes do not recognize l-nucleotides, spiegelmers can be made only by chemical synthesis, which limits access to longer spiegelmers. Ribozyme-catalysed cross-handed polymerization might enable convenient enzymatic access to spiegelmers, and eventually render them directly amenable to in vitro selection methods. Sczepanski and Joyce's twin polymerases will probably require further engineering before they can copy long RNA templates of any sequence efficiently and accurately. Nevertheless , successive improvements have been made for other in vitro-selected ribozymes 8,9 , providing reason for optimism in this case. ■