Steps at which eukaryotic gene expression can be controlled

Cell 7.5

Protein Variabilityand

Protein ActivityControl

Aminoacid sequence

Three-dimensional shape

(conformation)

Function

Protein processing

Degradation

Four types of post-translational processingevents

Protein Folding

The aminoacid sequence contains all the

information needed to fold the polypeptide into its

correct tertiary structure

The cellular mechanisms that monitor protein quality after protein synthesis

Cell 6.85

The co-translational folding of a protein

Cell 6.81

The cellular chaperone machinery is specifically recruited to bind to ribosomes and protects nascent chains and folding intermediatesfrom nonproductive interactions

Two main ATP-dependent classes of chaperons, Hsp70 chaperon and cylindrical chaperonin complexes, mediate protein folding

Structure of GroEL/GroES chaperonin(Hsp60)

Hsp70 chaperones bind tohydrophobic regions in unfolded polypeptides, including those thatare still being translated, and holdprotein in an open conformationuntil it is ready to be folded

Protein processing

Proteolytic Cleavage

Chemical Modification

(Intein splicing)

Protein processing by proteolyticcleavage

Genomes 11.27

Ex. proteolytic cleavage: melitin and insulin

Genomes 11.28

Promelittin Melitin

24 aa

22 aa

Extracellular protease

Signal peptide- hydrophobic aa sequence thatattachs preproinsulin to the membrane, beforeexporting the protein through the membrane tothe extracellular environment

A chain C chain B chain110 aa

30 aa21 aa

(51 peptide)

Converting preproinsulin to insulin

Ex. proteolytic cleavage : the pro-opiomelanocortinpolyprotein

POMC is glycosylated and then cleaved to give a numberof neurohormones that can be cleaved enzymaticallyinto the following peptides (neurohormones)

In anterior pituitary gland

In the intermediate lobeof pituitary gland

In the intermediate lobeof pituitary gland- melanin productionandneurons in the arcuatenucleus the hyphotalamus- apetite

Each of these peptides is packaged in large dense-corevesicles that are released from the cells by exocytosisin response to appropriate stimulation

260 amino acid protein

Chemical Modification

Post-translational chemical modificationof calf histone H3

Genomes 11.30

Lysine acetylation and methylation of histone H3

Various carboxylase enzymesLysineBiotinylation

Addition of biotin

Some protein kinases involved in signal transduction

N-terminal glycineN-myristoylation (miristic acid)

Many membrane proteinsSerine, threonine, cysteineAcylation

Addition of lipid side chains

Many membrane proteins and secreted proteins

AsparagineN-linked glycosylation (amino group)

Many membrane proteins and secreted proteins

Serine, threonineO-linked glycosylation (hydroxil groups)

Addition of sugar side chains

MelittinN-terminal glycineN-formylation (COH)

CollagenProline, lysineHydroxylation (OH)

Some proteins involved in signal transduction

Serine, threonine, tyrosinePhosphorylation (P)

HistonesLysineMethylation (CH3)

HistonesLysineAcetylation (CH3CooH)

Addition of small chemical groups

Examples of proteinsAmino acids that are modified

Modification

150 different aa already described

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 communicatedacross the plasma membrane to the gene regulatory proteins in the cell- SIGNAL TRANSDUCTION

Protein Degradation

Cell 6.82

Protein degradation... when?

Relation between N-terminal amino acid and half-life of E. coli ββββ-galactosidase proteins with modified N-terminal amino acids

N-terminal Amino Acid Half-life

Met, Ser, Ala, Thr, Val, Gly

Ile, Glu

Tyr, Gln

Pro

Phe, Leu, Asp, Lys

Arg

PESTsequence

ProGlnSerThr

Half-life less then 2 hours

more than 20 h

30 min

10 min

7 min

3 min

2 min

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

Lodish 3.13

Ubiquitin and the marking of proteinwith multiubiquitin chains

Ubiquitin-mediatedproteolytic pathway

(7-8 residues)