Microbiology 3-2

mRNA that often bears coding information transcribed from adjacent genes.

• Core Enzyme: 5 polypeptides
• Sigma Factor: Recognizes start of gene
• Holoenzyme= Core+Sigma

• The promoter is bound by RNA poly
• Sigma factor recognizes two regions in the promoter: -10 Pribnow box and -35 site.
• These are consensus sequences.

• RNA poly unwinds the DNA.
• Transcription bubble produced and moves with the RNA poly as it transcribes mRNA from template strand.

• Core RNA poly dissociates from template DNA
• 2 Types of termination: Intrinsic(AU rich region makes it fall off), and Rho-dependent(rho factor protein) reaches RNA poly and makes it pop off.

• Similar to bacteria: 1RNA poly & polycistronic mRNA
• Similar to eukarya: Promoter w/ TATA box, several transcription factors, some RNA has introns.

Up to 6 different codons for 1 amino acid

• Sense-Direct amino acid incorporation into the protein. (They code for actual amino acids.)
• Nonsense-The stop codons, involved in termination of translation.

Loose base pairing on the 5' end of tRNA, used to decrease mutation effects and reduce tRNA production needed.

• Structure:Cloverleaf, diamond bases, 2 arms, one acceptor stem and anticodon.
• Charging-Done through Aminoacyl-tRNA synthetase.  Attaches AA to 3' hydroxyl. 20 enzymes for each AA,

Ribosome serves as the binding site of the mRNA, where the proteins are put together (through linking AAs).

• 70S ribosome= 30S +50S subunits.
• 30S-16S rRNA & 21 proteins
• 50S- 5S and 23S rRNA & 34 proteins

• P-Peptidyl Site=Binds initiator tRNA or tRNA attached to growing polypeptide (peptidyl-tRNA)
• A-Aminoacyl site (Acceptor Site)=Binds incoming aminoacyl-tRNA
• E-Exit site=Briefly binds empty tRNA before it leaves ribosome

Involves: Ribosomal subunits, Initiation Factors (IFs), and Initiator tRNA w/ N-formyl methionine.

• Steps:fMet (N-formyl Methionine) binds to 30S.
• 30S aligns with Shine Delgarno seq and fMet binds with start/initiator codon.
• 50S binds to 30S w/ fMet at P site.
• IF3 prevents 30S binding to 50S prematurely (i.e. without mRNA).
• IF2 binds GTP to fMet and directs it to the P site of 30S.
• IF1 Helps release of IF2 and aids in binding of 50S and 30S.

• Aminoacyl tRNA binds
• Transpeptidation reaction occurs
• Translocation: Peptidyl tRNA moves from A site to P site, Ribosome moves down 1 codon, Empty tRNA leaves P site.

• EFs:
• EFTu-delivers aminoacyl-tRNA to the site.
• EFT-Converts EFTu-GDP back to EFTu-GTP.
• EFG-Helps with translocation by carrying GTP and GDP (moving the ribosome)

• Ribosome reaches the Nonsense codon (UAA, UAG, UGA) and it occupies the A site
• Release factors aid in recognition of stop (There are 3 RFs in procaryotes)

Eukary- Domains fold independently right after synth. Molecular chaperones=not as important

Prokary-Polypeptide doesn't fold till after entire synth of polypeptide. Moleculare chaperones are important.

Chaperones protect cells from thermal damage and aid in transport Xcross membranes. They also help proper folding

• Inteins-removed portion
• Exteins- portions that remain
• Splicing-Polypeptide is removed before final shape.

• Translocates proteins from cyto across or into plasma membrane.
• Secreted proteins synthed as preproteins having amino-terminal signal peptide
• -Peptide delays protein folding, chaperone proteins keep preproteins unfolded.

• From cytoplasm across plasma and outer membrane
• Secreted proteins have C-terminal secretion signals
• Gram+ bacteria use modified version of this pathway.

• 5 Secretion pathways
• II and V across outer membrane (Sec Dependent)
• I, III, IV Sec-independent

• II-From periplasmic space across outer membrane
• V-Moves proteins across outer membrane, autotransporters: form channel themselves. the Proteins make their own channels.

• III-Secretion of virulence factors in Gram - Bacteria
• From cytoplasm to outside of cell (some type III machinery is syringe shaped)
• IV- Secrete proteins and transfers DNA during Conjucation