COURSE CONTENT:
1. NUCLEIC ACIDS STRUCTURE AND TOPOLOGY : double-helical structure of DNA; different three-dimensional forms (A, B, Z); unusual structures; denaturation of double-helical DNA and RNA; hybrids; DNA supercoiling and topological properties; topoisomerases; structural organization of eukaryotic DNA ; nucleosomes and structure of chromatin.
2. DNA REPLICATION: structure and function of DNA-polymerases and other enzymes and proteins; mechanism of phosphodiester bond formation; processivity and proofreading activity of polymerases; replication of eukaryotic DNA, eukaryotic DNA-polymerases; replication of telomers, telomerases.
3. TRANSCRIPTION: initiation, elongation and termination of bacterial RNA synthesis; structure and function of bacterial RNA-polymerase, catalytic mechanism; eukaryotic transcriptional signals and diversity of RNA-polymerases; transcriptional factors.
4. POSTTRANSCRIPTIONAL EDITING and modifications of tRNA and rRNA transcripts (in bacteria and eukaryotes). RNase P as an example of the enzyme with RNA component. Posttranscriptional editing of eukaryotic transcripts; removal of introns; self-splicing; structure and function of spliceosome; 5 capping and 3 polyadenylation of mRNA . RNA enzymes.
5. TRANSLATION: structure of ribosome, tRNA and rRNA; initiation, elongation and termination of protein biosynthesis; aminoacylation of tRNA and proofreading; genetic code interpretation; role of elongation factors; tRNA suppressors; incorporation selenocysteine as a non-standard (21st) amino acid; inhibition of protein synthesis.
6. REGULATION OF GENE EXPRESSION IN BACTERIA AND EUKARYOTES: negative regulation by repressors, positive regulation by CAP-cAMP; attenuation; activation of chromatin, positive regulation of promotors in eukaryotes, chromatin remodeling.
7. PROTEIN TARGETING: posttranslational and co-translational import; posttranslational modifications.
LABORATORY WORK: denaturation and fragmentation of nucleic acids, gel electrophoresis, simple protein purification, protein-nucleic acids complex formation and analysis by gel-retardation assay; tRNA-mediated nonsense suppression
LEARNING OUTCOMES:
1. to show knowledge and understanding of biochemical reactions in the biosynthesis of nucleic acids and proteins
2. to interpret structure and catalytic mechanisms of enzymes and macromolecular assemblies participating in the transmission and flow of genetic information
3. to conclude about the importance of macromolecular interactions during transmission and flow of genetic information
4. to evaluate the importance of fidelity and processivity of the enzymes and corrections of errors during synthesis of natural polymers
5. to show knowledge and understanding of complex structure-function relationships between macromolecules participating in the transmission and flow of genetic information
6. to explain control mechanisms in the biosynthesis of nucleic acids and proteins with the emphasis on regulation of gene expression
7. to compare above mentioned biochemical processes and reactions in prokaryotes and eukaryotes
8. to independently perform experiments using standard biochemical methods and techniques
9. to appropriately handle biological macromolecules in order to avoid their denaturation and inactivation
10. to identify plasmids using restriction analysis
11. to determine melting curve and to calculate Tm of genomic DNA
12. to purify the protein using affinity chromatography
13. to isolate total tRNA from bacterial cells
14. to analyze macromolecular interactions between protein and nucleic acids using native electrophoresis (in vitro) and genetic methods (in vivo)
15. to write a report on laboratory excercises
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