Biochemistry of Nucleotides and mRNA Processing

Study Notes

Nucleosides are formed by a nitrogenous base attached to a sugar (ribose or deoxyribose). Examples include adenosine and deoxythymidine.
Nucleotides are nucleosides with one or more phosphate groups attached to the 5' carbon of the sugar.
Nucleoside di- and triphosphates are high-energy compounds due to the energy released from hydrolysis of their acid anhydride bonds. ATP is a key example.

Introns are removed from heterogeneous nuclear RNA (hnRNA) through splicing by spliceosomes (snRNPs).
Splicing occurs at the 5' (donor) and 3' (acceptor) splice sites of introns, excising them as a lariat structure.
Mutations in splice sites can cause abnormal proteins; mutations affecting β-globin mRNA splicing are implicated in some β-thalassemias.
The mature mRNA, after processing, is transported to the cytoplasm for translation.

Alternative splicing of primary transcripts produces multiple protein variants from a single gene.
This process is seen in muscle proteins (tropomyosin and troponin T) and immunoglobulin synthesis.
A large percentage of genes undergo alternative splicing, potentially contributing to the discrepancy between the number of genes and proteins in an organism (estimated ~30,000 genes vs. ~100,000 proteins in humans). These numbers are estimates and subject to change.
Alternative splicing can be detected using Northern blot analysis.

Ribosomal RNA (rRNA) is a component of ribosomes. Eukaryotic rRNA is transcribed by RNA polymerase I (45S precursor cleaved into 28S, 18S, and 5.8S rRNA) and RNA polymerase III (5S rRNA).
Eukaryotic ribosomes are 80S (60S and 40S subunits), while prokaryotic ribosomes are 70S (50S and 30S subunits). The "S" values represent sedimentation coefficients and are not additive.
Transfer RNA (tRNA) carries activated amino acids during translation. Eukaryotic tRNA is transcribed by RNA polymerase III. Each tRNA carries a specific amino acid. tRNAs share a general cloverleaf structure but have distinct features.

Gene regions: Prokaryotic genes are often polycistronic (multiple genes in one mRNA), while eukaryotic genes are monocistronic (one gene per mRNA). Eukaryotic genes contain exons (coding sequences) and introns (non-coding sequences).
RNA polymerase: Prokaryotes have a single RNA polymerase; eukaryotes have three: RNA polymerase I (rRNA), RNA polymerase II (mRNA, snRNA), and RNA polymerase III (tRNA, 5S rRNA).
Transcription initiation: Prokaryotes use a sigma factor to recognize the promoter region (-35 sequence and -10 sequence (Pribnow box, TATAAT)); eukaryotes use transcription factors such as TFIID which binds to the TATA box (-25 sequence) and CAAT box (-70 sequence).
Transcription termination: Prokaryotes employ stem-loop structures and often a rho factor; eukaryotic termination is less well-defined.
mRNA processing: Prokaryotic mRNA does not undergo significant processing; eukaryotic mRNA acquires a 5' cap (7-methylguanosine), a 3' poly(A) tail, and undergoes intron removal (splicing).
Ribosomes: Prokaryotic ribosomes are 70S, while eukaryotic ribosomes are 80S.
tRNA: tRNAs share a cloverleaf secondary structure with an acceptor arm (CCA) for amino acid attachment and an anticodon arm complementary to mRNA codons.