Biochemistry - The Chemical Reactions of Living Cells, Volume 3 - D. Metzler 1980

Biochemical Genetics and the Synthesis of Nucleic Acids and Proteins
How the Current Concept Developed
Ribonucleic Acids and Proteins

By 1942, data obtained using the ultraviolet cytophotometry method developed by Caspersson [30], along with the results of Brachet's cytochemical studies [31], made it clear that RNA is somehow involved in Protein Synthesis. Autoradiographic studies using ³H-uridine showed that in Eukaryotic Cells, RNA is synthesized in the nuclei and subsequently transported to the Cytoplasm [32, 33]. The existence of Ribosomes was discovered by electron microscopists studying The Structure of the cytoplasmic Endoplasmic reticulum using ultrathin sections. Their presence in The Cell was firmly established in 1956, and the term ribosome was proposed in 1957. Over several years, isolated ribosomes became the focus of intensive research aimed at elucidating The Mechanism of METABOLISM/35.html">Protein Biosynthesis. Initially, studying Protein synthesis in vitro proved quite challenging because there was no method yet for quantifying newly synthesized protein. However, Hoagland et al. [33a] developed such a method. Their approach was based on measuring The rate of incorporation of ¹⁴C-labeled Amino Acids into Proteins. This extremely sensitive method made it possible to measure minute quantities of protein synthesized in cell-free rat Liver preparations and paved the way for studying protein synthesis on the ribosomes themselves.

Immediately after the appearance of the Watson and Crick hypothesis in 1953, it was suggested that ribosomal RNA (rRNA), which accounts for up to 90% of total cellular RNA in some cells, serves as the carrier of Genetic information from The Nucleus to the cytoplasm. However, by 1960 it was shown that this assumption was incorrect. Specifically, despite significant differences in the Nucleotide Composition of DNA, the size and nucleotide composition of RNA from the ribosomes of various Bacteria were found to be very similar (Chap. 2, Sec. D, 8) [34]. Furthermore, by that time it became clear that information transfer is mediated by a relatively unstable, short-lived form of RNA, whereas ribosomal RNA proved to be highly stable [35].

a. Messenger RNA (mRNA)

Evidence for the existence of a labile form of RNA was obtained in 1956 by Volkin and Astrachan [35a], who discovered rapidly labeled RNA in phage-infected bacterial cells. Enzyme induction studies (Chap. 6, Sec. E, 2) also played a crucial role in the discovery of mRNA, demonstrating that many bacteria, including E. coli grown on a medium with glucose as the sole energy source, do not immediately acquire The ability to utilize a new sugar when transferred to a lactose medium. However, within 2 minutes of transfer to the lactose medium, they begin to synthesize new proteins required for lactose metabolism. These new proteins include Enzymes such as lactose permease and ß-galactosidase, which splits the disaccharide into glucose and galactose. When the lactose supply in the medium is exhausted, The activity of the inducible enzymes drops almost as rapidly. These findings indicated that the RNA carrying the genetic information for the synthesis of new enzymes is seemingly unstable; it must be synthesized rapidly in response to the appearance of an inducing sugar and disappear quickly in its absence.

In 1961, Jacob and Monod [36] postulated the existence of a short-lived messenger RNA (mRNA). By that time, there were numerous other arguments supporting this assumption. It was found, for example, that RNA molecules produced following the infection of E. coli with bacteriophage T4 hybridize (Chap. 2, Sec. D, 10) with denatured bacteriophage DNA. Moreover, this virus-specific mRNA was shown to associate with pre-existing bacterial ribosomes and serve as a template for the synthesis of phage proteins [37]. This experiment directly pointed to the Transcription of mRNA from viral DNA genes.

b. Transfer RNA

In 1957, Crick [37a] suggested that special "adaptor" molecules are required to arrange amino acids in an order corresponding to The sequence of their codons in the transcribed RNA. Crick believed that these adaptors might be polynucleotides. By that time, chemical studies of cellular RNA had revealed that low-molecular-weight RNA accounts for up to 15% of the total RNA in cells. In the same year (1957), Hoagland's Discovery of the enzymatic "activation" of amino acids prior to their incorporation into proteins suggested that these very low-molecular-weight RNA molecules act as the postulated adaptors. It was proposed to call this RNA transfer RNA (tRNA, Fig. 2-24).

In recent years, an "army" of dedicated researchers in protein synthesis has greatly expanded our knowledge on the subject. Now, however, it seems more appropriate to move on from tracing the historical events associated with The problem of protein synthesis to examining some of the details, starting with RNA transcription—a process that has been studied in considerable detail.