Gene Action and Amino Acid Coding 



433 



amino acid meaning in the RNA code, since 

 regardless of the length of the polyribotides 

 of A or C added, no amino acid is incor- 

 porated into protein. But even if each single 

 ribonucleotide specified a different amino 

 acid, only four amino acids would be en- 

 coded. But 20 amino acids need be en- 

 coded! Can we circumvent this difficulty by 

 assuming that a sequence of two nucleotides 

 specifies an amino acid? (This would be 

 like having an alphabet of only four letters 

 and a language composed only of two-letter 

 words.) In this case, since the first letter can 

 be any one of four, as can the second, there 

 are 4x4, or 16, different doublets (or 

 words) possible, if the presumption is made 

 that the RNA code can be read only in one 

 direction. Unidirectional reading seems 

 reasonable, since a single strand of RNA is 

 polarized, just as is a single strand of DNA. 

 Still, this will not be enough to specify 

 20 amino acids, unless other assumptions are 

 made. (For example, it could be hypothe- 

 sized that a given doublet encodes more 

 than one kind of amino acid, in which case 

 we would be dealing with a degenerate code.) 

 If, however, a sequence of three nucleotides, 

 that is, a triplet, encodes an amino acid, there 

 would be 4x4x4, or 64, different uni- 

 directional sequences possible. This pro- 

 vides more than enough triplets to encode 

 20 amino acids. (The code would also be 

 degenerate if different triplets encoded the 

 same amino acid.) 



There are, however, other characteristics 

 of RNA which have a bearing on the nature 

 of its code. Since the number of consecutive 

 ribotides may be in the hundreds or thou- 

 sands, there are no spaces (or non-nucleotide 

 punctuation) to indicate where one triplet 

 should stop and the next should begin. Ac- 

 cordingly, we are dealing with what may be 

 called a comma-free code. Suppose we had 

 six ribotides arranged linearly, in positions 

 called 123456. If triplet 123 specifies amino 

 acid A and 456 amino acid B, errors would 



be possible if the overlapping triplets 234 or 

 345 served to encode different amino acids, 

 and if the reading of different meaningful 

 triplets were performed independently. In 

 this event, the last two triplets would have to 

 be eliminated as words in the code to avoid 

 errors. Accordingly, the entire code has to 

 be examined, and all overlapping triplets 

 eliminated from being meaningful, that is, 

 from specifying an amino acid. When, of 

 the 64 possible triplets, all overlapping ones 

 are eliminated, 20 nonoverlapping triplets 

 (amino acid-specifying "words") remain. 

 However, the question of overlapping triplets 

 can be avoided to a great extent if the read- 

 ing of the message can start only at a given 

 place and triplets are read in succession. In 

 this case, the punctuation is provided by the 

 mechanism for reading the code. The least 

 we can conclude from the present discussion 

 is that it requires a sequence of more than 

 one ribotide to encode an amino acid. 



When the synthetic polyribotide of U is 

 mixed with the synthetic polyribotide of A, 

 so that several strands are likely to base pair 

 or wrap about one another, incorporation of 

 phenylalanine is partially or completely re- 

 duced. Thus, the synthetic polymer is most 

 effective in synthesis when it is single- 

 stranded, as is apparently also true for nor- 

 mal messenger RNA. 



It is also possible to study the effect on 

 amino acid incorporation into protein of the 

 presence of different nucleotides in the same 

 polyribonucleotide. Using polynucleotide 

 phosphorylase, one can enzymatically syn- 

 thesize in vitro polyribotides containing two 

 or more different ribotides which are ap- 

 parently in a random order. The analy- 

 sis''^ is greatly expedited by the fact that 

 polyphenylalanine is insoluble in the cell- 

 free system. In practice, then, in forming 

 any mixed polynucleotide, an excess of 

 uridylic acid is used, in order to obtain the 

 synthesized protein as a precipitate, which 

 ' By S. Ochoa and coworkers. 



