The Reading Frames

Frameshift Mutation A type of mutation in which a number of nucleotides not divisible by three is deleted from or inserted into a coding sequence, thereby causing an alteration in the reading frame of the entire sequence downstream of the mutation. These mutations may be induced by certain types of mutagens or may occur spontaneously. [NIH]... 

In-frame mutation A mutation, usually a deletion, which does not shift the reading frame downstream of the mutation. 

Frameshift mutations (choice A) are the result of the deletion or insertion of a series of nucleotides that are not a multiple of three (thus altering the reading frame). Although the insertion or deletion of a single nucleotide would produce a frameshift, it is highly unlikely that it would alter only a single amino acid. The shift in the reading frame typically alters a number of amino acids subsequent to the insertion or deletion site. 

Nitrous acid causes point mutations (1). For example, C is converted to U, which in the next replication pairs with A instead of G. The alteration thus becomes permanent. Mutations in which a number of nucleotides not divisible by three are inserted or removed lead to reading errors in whole segments of DNA, as they move the reading frame (frame-shift mutations). This is shown in Fig. 2 using a simple example. From the inserted C onwards, the resulting mRNA is interpreted differently during translation, producing a completely new protein sequence. 

The cytoplasmic organelles that provide the platform for this process are the ribosomes. This complex recognizes and binds to a specific region at the 5 end of the mRNA. The signal for initiating a polypeptide chain is a special initiation codon in the mRNA sequence that marks the start of the reading frame. Usually the initiation codon is the triplet AUG, which represents the amino acid methionine, and thus most proteins have this amino acid as the first one in the polypeptide. 

If the fusion of a tag is planned, it is important to consider the reading frames of an ORF and the tag so that they are properly translated in-frame. In addition, removal of the termination codon is essential for production of C-terminally tagged proteins. 

Frame-Shift Mutation A change in the structure of DNA such that the transcription of genetic information into RNA is completely altered because the start point for reading has been altered i.e. the reading frame has been altered. 

Detection of intron/exon splice sites Introns in eukaryotes cause discontinuation of the reading frame. If the analysis is not focused on a cDNA sequence, these introns must be spliced out and the exons joined to form the sequence that actually codes for the protein. Intron/exon splice sites can be predicted based on their common features. Most introns begin with the nucleotides GT and end with the nucleotides AG. There is a branch sequence near the downstream end of each intron involved in the splicing event. 

One of the best-documented examples occurs in translation of the mRNA for the overlapping gag and pol genes of the Rous sarcoma virus (see Fig. 26-31). The reading frame for pol is offset to the left by one base pair (—1 reading frame) relative to the reading frame for gag (Fig. 1). 

Messenger RNA Translation Predict the amino acid sequences of peptides formed by ribosomes in response to the following mRNA sequences, assuming that the reading frame begins with the first three bases in each sequence. 

Can the Base Sequence of an mRNA Be Predicted from the Amino Acid Sequence of Its Polypeptide Product A given sequence of bases in an mRNA will code for one and only one sequence of amino acids in a polypeptide, if the reading frame is specified. From a given sequence of amino acid residues in a protein such as cytochrome c, can we predict the base sequence of the unique mRNA that coded it Give reasons for your answer. 

Frame-shift mutations If one or two nucleotides are either deleted from or added to the interior of a message sequence, a frame-shift mutation occurs and the reading frame is altered. The resulting amino acid sequence may become radically different from this point on (Figure 31.5). [Note If three nucleotides are added, a new amino acid is added to the peptide or, if three nucleotides are deleted, an amino acid is lost. In these instances, the reading frame is not affected.]... 

Frame-shift mutations as a result of addition or deletion of a base can cause an alteration in the reading frame of mRNA. 

Messenger RNA is read by the protein synthesizing machinery from some starting point. As is illustrated in Chapter 5, Section E,1 the codons are read three bases at a time, and the proper amino acid corresponding to each codon is inserted. When a deletion or insertion in the mRNA is met, all subsequent codons may be misread because the reading frame has shifted forward or backward by one or two nucleotides. The protein synthesized bears little resemblance to that formed by the nonmutant organism and is usually completely nonfunctional. Mutations are considered further in Chapter 27. 

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