Steps at which eukaryotic gene expression can be controlled. Cell 7.5

Transcription

1 Steps at which eukaryotic gene expression can be controlled Cell 7.5

2 Protein Variability and Protein Activity Control Aminoacid sequence Three-dimensional shape (conformation) Function Protein processing Degradation

3 Four types of post-translational processing events

4 Protein Folding The aminoacid sequence contains all the information needed to fold the polypeptide into its correct tertiary structure

5 The cellular mechanisms that monitor protein quality after protein synthesis Cell 6.85

6 The co-translational folding of a protein The cellular chaperone machinery is specifically recruited to bind to ribosomes and protects nascent chains and folding intermediates from nonproductive interactions Cell 6.81

7 Two main ATP-dependent classes of chaperons, Hsp70 chaperon and cylindrical chaperonin complexes, mediate protein folding Hsp70 chaperones bind to hydrophobic regions in unfolded polypeptides, including those that are still being translated, and hold protein in an open conformation until it is ready to be folded Structure of GroEL/GroES chaperonin (Hsp60)

8 Protein processing Proteolytic Cleavage Chemical Modification (Intein splicing)

9 Protein processing by proteolytic cleavage Genomes 11.27

10 Ex. proteolytic cleavage: melitin and insulin Promelittin Melitin 22 aa Extracellular protease 24 aa A chain C chain B chain 21 aa 30 aa 110 aa Signal peptide- hydrophobic aa sequence that attachs preproinsulin to the membrane, before exporting the protein through the membrane to the extracellular environment Genomes 11.28

11 Converting preproinsulin to insulin (51 peptide)

12 Ex. proteolytic cleavage : the pro-opiomelanocortin polyprotein 260 amino acid protein POMC is glycosylated and then cleaved to give a number of neurohormones that can be cleaved enzymatically into the following peptides (neurohormones) In anterior pituitary gland In the intermediate lobe of pituitary gland In the intermediate lobe of pituitary gland- melanin production and neurons in the arcuate nucleus the hyphotalamus- apetite Each of these peptides is packaged in large dense-core vesicles that are released from the cells by exocytosis in response to appropriate stimulation

13 Chemical Modification

14 Post-translational chemical modification of calf histone H3 Lysine acetylation and methylation of histone H3 Genomes 11.30

15 Modification Amino acids that are modified Examples of proteins Addition of small chemical groups Acetylation (CH 3 CooH) Methylation (CH 3 ) Phosphorylation (P) Hydroxylation (OH) N-formylation (COH) Lysine Lysine Serine, threonine, tyrosine Proline, lysine N-terminal glycine Histones Histones Some proteins involved in signal transduction Collagen Melittin Addition of sugar side chains O-linked glycosylation (hydroxil groups) N-linked glycosylation (amino group) Addition of lipid side chains Acylation N-myristoylation (miristic acid) Addition of biotin Biotinylation Serine, threonine Asparagine Serine, threonine, cysteine N-terminal glycine Lysine Many membrane proteins and secreted proteins Many membrane proteins and secreted proteins Many membrane proteins Some protein kinases involved in signal transduction Various carboxylase enzymes 150 different aa already described

16 Some ways in which the activity of gene regulatory proteins is regulated in eucaryotic cells Each of these mechanisms is typically controlled by extracellular signals which are communicated across the plasma membrane to the gene regulatory proteins in the cell- SIGNAL TRANSDUCTION

17 Protein Degradation

18 Protein degradation... when? Cell 6.82

19 Relation between N-terminal amino acid and half-life of E. coli β- galactosidase proteins with modified N-terminal amino acids N-terminal Amino Acid Met, Ser, Ala, Thr, Val, Gly Ile, Glu Tyr, Gln Pro Phe, Leu, Asp, Lys Arg Half-life more than 20 h 30 min 10 min 7 min 3 min 2 min PEST sequence Pro Gln Ser Thr Half-life less then 2 hours

20 Acerca da degradação de proteínas Várias vias de degradação Lisossomas: contêm uma série de hidrolases e enzimas proteolíticos, degradando essencialmente proteínas transmembranares e do lúmen dos organitos Proteossoma: complexo multiproteico que degrada proteínas ubiquitinadas, localizadas sobretudo no núcleo e no citosol. Ex. Factores de transcrição, proteínas da regulação do ciclo celular como as cinases e fosfatases etc. - Processos altamente selectivos e rápidos Eucariotas- descrito o Proteossoma

21 Ubiquitin and the marking of protein with multiubiquitin chains Ubiquitin-mediated proteolytic pathway (7-8 residues) Lodish 3.13