Twelve years ago I wrote an Alternate View column (Analog, October, 1989) reporting on Nanocon I, a conference that had just been held in Seattle on February 17-19, 1989. Nanocon I, which I helped to organize, was the world’s first technical and scientific conference devoted specifically to the emerging field of nanotechnology. Nanotechnology gets its name from the nanometer, a distance of 10-9 meters or roughly the diameter of a molecule, and the term refers to the technology for structuring matter with precise control at the nanometer scale, atom-by-atom or molecule-by-molecule, to form a pre-specified pattern. In other words, nanotechnology is the general ability to build large or small structures to complex atomic specifications. In 1989 nanotechnology was a relatively unfamiliar concept in science fiction. In the past decade it has exploded to become a major theme in of much science fiction, including my own novel Einstein’s Bridge.
However, while there have been some impressive technical achievements in this area in the past decade, one of the major goals discusses extensively at Nanocon I has not yet been realized. That goal is the production of an assembler, a device that under programmatic control can assemble atoms and molecules to produce nanomachines, including another assembler. Can we be sure that goal is actually possible? Yes, because wee have what mathematicians call an existence theorem. Biological systems as simple as bacteria are already practicing nanotechnology by constructing proteins using an assembler of sorts, the ribosome.
In this column, I want to describe the recent accomplishment of a major step on the path to producing a general assembler, the mapping of the atomic structure of the ribosome, Nature’s own assembler of proteins. We’ll start by describing in some detail how the ribosome assembles a protein. The inner mechanisms of life are rather complicated, so buckle your seatbelts!
Proteins are strings of amino acids that self-fold into units that are the intricate building blocks of all living things. Proteins form the structural members, regulators, defenders, catalysts, communicators, pumps, movers, and shakers of living organisms. Each protein is assembled by a ribosome using a set of instructions encoded in a strand of messenger RNA. The messenger RNA (mRNA) is a working copy of the information transcribed from the DNA “central library” of a cell, but it has a slightly different form. DNA has a backbone of deoxyribose and phosphate supporting an alternating sequence of the nucleotides thymidylic acid (T), cytidylic acid (C), adenylic acid (A), and guanylic acid (G), while mRNA has a backbone of ribose and phosphate supporting an alternating sequence that has the nucleotide uridicylic acid (U) substituted for thymidylic acid (T) but otherwise uses the same C, A, and G nucleotides found in DNA. The structural differences make mRNA a more mobile linear sequence that does not have the DNA tendency to form a double helix. The linear nucleotide structure of mRNA is read like a strip of punched paper tape, three nucleotides at a time, by the ribosome as the protein is constructed.
The sequence of mRNA instructions for assembling a protein can be thought of as a long “sentence” constructed from a string of “words, each consisting of three “letters” drawn from a four letter “alphabet”, with the sentence ultimately terminated by a “period” or stop command.. The alphabet is the four nucleotides U, C, A, and G. In principle, there should be 43 or 64 different three-letter combinations forming words constructed from an alphabet of four letters. Three of the possible three-letter combinations, UAA, UAG, and UGA, are stop commands that inform the ribosome that the protein sequence is completed and that assembly should halt. Many of the other combinations are redundant, with several different three-letter codes specifying the same amino acid. For example, the three-letter sequences UUA, UUG, CUU, CUC, CUA, and CUG are all instructions to attach the same amino acid, leucine. Because of these redundancies the 61 possible three-letter combinations that are not stop commands actually code for only different 20 amino acids. Those 20 amino acids specified by the mRNA code are lysine, argenine, histidine, aspartic acid, glutamic acid, asparagines, glutamine, serine, threonine, tyrosine, glycine, alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan, and cysteine. It is interesting to note that there are other natural amino acids that are not on this list of 20 and therefore cannot be included in protein synthesis by ribosomes.
The ribosome does not directly collect and assemble amino acids. It has help from special transfer RNA (tRNA) molecules, rather short RNA strands that are the amino acid handlers in the protein synthesis process. The tRNA molecule has one special section that contains a complementary sequence that matches and docks with the three-letter mRNA code just described. At one end of this tRNA molecule is attached an activated form of the amino acid to which the three-letter code refers. These tRNA molecules collect amino acids, transport them to the ribosome, and participate in assembling them into a protein. At the end of this process, the used tRNA, now lacking its amino acid, is released. It then finds its way to a special protein enzyme that loads it with another activated amino acid so that the process can repeat.
The ribosome itself is a special molecule that comes in two pieces. The smaller piece is called the 30S subunit, and the larger piece the 50S subunit. Here, the S designations come from the sedimentation rate of the molecule in a centrifuge, which is related to the molecule’s size and weight. In shape the 30S subunit is rather like the handset of an old-style 1970s telephone and the 50S subunit is like the telephone’s base. The protein synthesis begins when the 30S attaches to the beginning of a strand of mRNA. Then an initial molecule of tRNA having coding that matches the first word of the mRMA strand attaches to the first or “A” site of the 30S subunit. Then the 30S and 50S subunits close together, like hanging up the telephone, trapping the mRNA strand in the ribosome’s reading mechanism so that protein synthesis can begin.
The two subunits of the ribosome, now acting together, shift the mRNA and attached tRNA to the second or “P” site of the 30S subunit and allow a second tRNA molecule to doc with the three-letter mRNA pattern now at the A site. There is believed to be a “proof-reading” process in which the match between the mRNA and their complements in the tRNA unit is verified. The amino acid at the end of the tRNA molecule at the P site is joined to the amino acid of the new tRNA molecule at the A site and deactivated, forming the first link of the protein’s backbone. Then the ribosome shifts both tRNA molecules over one position, so that the first tRNA molecule moves to the “E” or exits site of the 30S subunit . There the first mRNA molecule, now detached from its amino acid, is released to be recycled. A new tRNA molecule docks at the A site of the ribosome and the process continues. The protein grows link by link as the mRNA strand is processed through the ribosome until a stop code is reached. When this happens, the ribosome opens, like lifting the telephone handset from the base, and the protein and last used tRNA molecule are released. Since the mRNA strands are often quite long, they may be sequentially read by many ribosomes in a row, with protein synthesis occurring in parallel in all of the ribosomes involved.
It is now clear that the ribosome is not a purely protein biological structure. About 2/3 of the ribosome’s mass is composed of RNA with the other 1/3 made up of protein components. Further it is now clear that most of the essential functions of the ribosome are carried out by the RNA components.
The two components of the ribosome are very different. The 30S unit does the decoding and mRNA transport while the 50S unit is the catalyzer that, when properly triggered by her 30S subunit, actually produces the protein. The 30S unit has a unique structure, with clear domain boundaries and flexible regions capable of the large movements needed to move the mRNA strand through the ribosome. In contrast, the 50S subunit is relatively solid and monolithic, its components intricately folded and locked together into a rigid structure.
The decoding of the ribosome has proved difficult because the subunits are so large. The standard method for such mapping is to collect a large quantity of the molecules of interest and then form a repetitive crystal structure that can be studied using the technique of x-ray diffraction. The 50S subunit had already been crystallized and mapped by x-ray diffraction to a spatial resolution of 0.24 nanometers. Now a group of molecular biologists from Camdridge, UK, Salt Lake City, Utah, and Göttingen, Germany has been able to use the technique to map the structure of the 30S subunit to a spatial resolution of 0.3 nanometers, thereby producing a complete description of the 30S structure. Their paper in the journal Nature includes elaborate stereoscopic views of the structure and its substructures. The same group in a separate paper also studied crystals formed with the 30S subunit combined with certain antibiotic molecules that are known to interfere with protein synthesis in the ribosome. This promises to provide new insights into how antibiotics work and how they may be improved.
The process of protein synthesis described above indicates that the ribosome, the cell’s protein factory, is a complex and intricate bio-machine. It is a fundamental component of life forms from bacteria to multicellular organisms, plants, and animals. It is a key element of life itself. We would like to understand precisely this keystone of biological processes works. The first step in this process, the mapping of the structure of the ribosome, has been accomplished. The next step is to understand the map, and to reach an understanding of how the ribosome works. This will involve detailed computer modeling and testing.
We would also like to discover how the ribosome can be put to other uses besides the fabrication of proteins, how it can be modified, improved, and extended. Our goal is to take over Nature’s assembler for our own purposes, put its great capabilities to our own uses. We are noth there yet, but we are getting closer.
Protein Synthesis and RNA:
The Way Life Works, Mahlon Hoagland and Bert Dodson, Times Books Division of Random House, New York (1998), ISBN 0-8129-2888-1.
Structure of the Ribosome:
N. Ban, et al., “Structure of the 50S ribosomal subunit”, Science 289, 905-920 (2000).
P. Nissen, et al., “Structure of the 50S ribosomal subunit”, Science 289, 920-930 (2000).
Brian T. Wimberly, et al, “Structure of the 30S ribosomal subunit”,Nature 407, 327 (2000).
Andrew P. Carter, et al, “Functional insights from the structure of the 30S ribosomal subunit and its interactions with antibiotics”,Nature 407, 327 (2000)
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