A membrane transporter for tryptophan composed of RNA

Abstract

We have incorporated an RNA binding site for the biological amino acid tryptophan within an RNA complex with affinity for phospholipid bilayer membranes. The resulting RNA (9:10 Trp ) creates a selective route through the bilayer for the amino acid. Binding and enhanced tryptophan permeability are nonlinear in RNA concentration, suggesting that RNA aggregation is required for both. Tryptophan permeability saturates with increased concentration, though at ∼ 1000-fold greater level than when binding a free aptamer. The RNA (9:10 Trp ) complex, bound at a mean of two per liposome, halves the activation energy for tryptophan transport (to 46 kJ/mole), specifically increasing tryptophan entry to a maximal velocity of 0.5 sec −1 per liposome with little or no accompanying increase in general permeability. Individual RNAs turn over tens of thousands of times at high tryptophan concentration. Thus, a specific passive membrane transporter whose properties overlap those of single-molecule transporter proteins, can be made of RNA alone. Permeability changes probably rely on disturbances in lipid conformation as well as on an advantageous low free energy position for tryptophan at the membrane. Other RNA activities may yield other RNA-membrane nanosystems via this route.

Membrane Transport in Primitive Cells

“…As some permeability mechanisms invoke transient pores and packing defects as routes for solute passage, it is plausible that RNA molecules that simply interact with membrane surfaces could influence the structure and dynamics of the membrane enough to effectively enhance solute permeation. In fact, a RNA chimera consisting of selected membrane and tryptophan binding motifs is able to facilitate tryptophan transport across diacyl phospholipid membranes (Janas and Yarus 2004). Although few examples exist of such RNAmembrane interactions, the ability of RNA to serve a wide range of functions in vitro and in contemporary cells suggests that facilitated solute transport may not have been impossibility during the origin of life ).…”

Synthetic biology projects in vitro

“…Impressively, ribozymes that polymerize nucleoside triphosphates have been constructed, but improving the fidelity and efficiency to the levels needed for replication remains a significant hurdle (Szostak et al 2001). Another hurdle is construction of a membrane transporter made of RNA despite its membrane impermeability, yet this has recently been proved feasible (Janas et al 2004). A benefit of this approach is experimental insight into the extinct "RNA world.…”

Human tRNASecassociates with HeLa membranes, cell lipid liposomes, and synthetic lipid bilayers

“…Thus, specific nucleotide sequences are required for enhanced affinity to phospholipid bilayers but such RNA domains, judging from ease of selection, must be small and numerous. By combining such preselected RNA-membrane affinity domains with an RNA amino acid binding RNA site, a passive membrane transporter specifically directed to the amino acid tryptophan was constructed and characterized (Janas et al 2004). This membrane RNA, composed only of the four standard ribonucleotides, is specific to the amino acid side chain and transports tryptophan at a rate that overlaps proteins carrying out a similar membrane transport reaction.…”