Post-transcriptional cleavage

The data of Figure 10 illustrate that post-synthetic modification, including hydrolysis, is not limited to proteins. In this example, the large pre-mRNA undergoes post-transcriptional cleavage, splicing, modification of the 3 and 5 ends, and finally methylation to give the specific active mRNA. 

A primary transcript is a linear copy of a transcriptional unit—the segment of DNA between specific initiation and termination sequences. The primary transcripts of both prokaryotic and eukaryotic tRNAs and rRNAs are post-transcriptionally modified by cleavage of the original transcripts by ribonucleases. tRNAs are then further modified to help give each species its unique identity. In contrast, prokaryotic mRNA is generally identical to its primary transcript, whereas eukaryotic nrRNA is extensively modified posttranscriptionally. 

Most mature mRNAs have a post-transcriptionally added poly (A) tract at their 3 end (Wahle and Ruegsegger, 1999). Usually preceding the poly (A) addition site is a sequence transcribed from the sequence AATAAA on the DNA. This sequence, the so-called poly (A) signal, is the most conserved recognition element in eukaryotes. This sequence motif directs the cleavage of primary transcripts... 

The 3 -end of an eukaryotic mRNA is formed post-transcriptionally by endonucleolytic cleavage downstream of the coding sequence followed by extension of the upstream fragment by approximately 200 adenylate residues. In higher eukaryotes the specificity of the reaction depends on two conserved sequences in the pre-mRNA a highly conserved AAUAAA 10-30 nucleotides upstream from the cleavage site and a G/U-rich sequence within 50 nucleotides downstream from this site. The reaction requires a complex set of nuclear proteins, and a reconstituted polyadenylation reaction from pure proteins has not yet been achieved. However, processing of... 

Transcription produces RNA molecules. In prokaryotes, rRNAs and tRNAs are synthesized as longer precursors which require post-transcriptional processing including methylation and cleavage by specific endonucleases before they are functional molecules. Polycistronic mRNAs (Section 7.4) do not require further enzymic modification. 

Very often, no exact database match can be found even with high quality MS/MS mass spectra. It depends directly on the completeness and accuracy of the database searched, i.e. whether the genome is complete or incomplete, and on the quality of the transcripted EST sequences. These problems raise e question whether it is a novel protein, a known protein with a post-translational modification or if the failure to produce a database match due to inter-species variation, database sequence errors, or unexpected proteolytic cleavages. To address this problem, de novo interpretation of MS/MS data is an alternative where the amino acid sequence of peptides is derived by interpreting the mass differences between the generated MS/MS fragment ions sequence. 

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