Post-translational Modifications to Proteins

Affecting Form and Function

Contents
 Why

are proteins modified?  Glycosylation  Membrane proteins  Proteolytic processing  Phosphorylation  Acetylation  Small Molecule Binding  Regulated degradation

Why are proteins modified?
 Regulation

of activity

modification may turn activity on  modification may turn activity off  modification may generate a different function
 Protein-protein

interaction

modification site may be a binding interface

 Subcellular

localization

modification site may be a targeting signal  modification may be a membrane anchor
 Aging

modification may identify the protein for degradation  modification may target a protein to be scavenged

Topics to cover in my section
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593-595

Membrane proteins - myristlyation, farnesylation

598,612-3,735-736 Glycosylation 702-704 Glycosylation in rough ER 706,763-764 GPI anchors 733-735 Oligosaccharide processing (Golgi & ER, Glyco-related) 742-745 M6P in lysosomal enzymes 1085-6 Lectins and the immune system 1091-6 Proteoglycans and extra-cellular matrix 678-681,685,694 Cleavage of signal sequences (mitochondria, chloroplasts, ER) 760 Proteolysis of secreted proteins (to activate them) 893-895 Notch cleavage in neuron development

Glycosylation
Major

form of protein modification Sugars are added in the ER and Golgi Most proteins formed in the ER are glycoproteins Many different forms and functions

Initial glycosylation in the ER
A

precursor oligosaccharide is formed on a dolichol lipid This is transferred to the growing protein

Glycosylation and protein folding

by glucosidase

Processing in the Golgi

Functions of glycosylation
Stabilise

proteins against proteolysis

Limit approach of macromolecules to protein surface

Modulation

of immune response

Selectins (weakly) bind to oligosaccharides  Helps to concentrate lymphocytes in lymphoid organs  Attracts white blood cells & platelets to inflammation sites

Functions of glycosylation
Provide

sorting signals

M6P for lysosomal hydrolases  GPI anchors (see later)
Contributes

to differentiation events in organism development
Removing N-acetylglucoasmine transferase I in mice causes embryo death
 Neural

tube development and left-right body plan asymmetry impaired

Proteoglycans and the Extracellular Matrix
Made

of core protein and polysaccharide chains
Extremely diverse

Form
 

hydrated gel

Resists compressive forces Regulate traffic (perlecan in kidney)

Can

regulate secreted protein activity
e.g. chemokines in inflammatory response

Membrane proteins: GPI anchors

GPI-anchored

proteins are delivered to the apical plasma membrane Trypanosomes can shed these proteins to avoid immune attack

Myristylation and Farnesylation
Attaches

cytosolic proteins to the plasma membrane Protein usually involved in signal transduction

Proteolytic processing

Why

is this common for secreted enzymes?
Some peptides (e.g. enkephalins) too short by themselves  Prevent premature activation of hydrolytic enzymes

Phosphorylation
Most common posttranslational modification to proteins in eukaryotes  Enzymes and regulators are turned ‘on’ and ‘off’  Energy from ATP

Phosphorylation Regulates Protein Synthesis – eIF-2

Phosphorylation and Molecular Switches

Signalling using GTP-Binding - Ras Protein
 Broadcasts
 Cell

signals from cell surface

proliferation  Differentiation

Phosphorylation and Motor Proteins
 Move

chromosomes during mitosis  Move organelles along molecular tracks  Move enzymes along DNA during DNA synthesis

Phosphorylation and Motor Proteins
 ATP

binding - conformation 1 to conformation 2  ATP hydrolyzed to ADP Pi conformation 2 to conformation 3.  Release of ADP and Pi back to conformation 1.  Irreversible – one direction only

Acetylation

Acetylation and Histones
 Acetylation

enhances transcription  Deacetylation represses transcription

Small Molecule Binding

Retinal B. Heme group
A.

Protein Degradation
 Degradation
 Ubiquitin

mechanisms:
ligase  Degradation signal

 Multiubiquitin

chain marks protein for degradation in proteosome

Common Post-translational Modifications
Sulphydryls Amines Disulphide bond Cysteinylation Methylation Acetylation Farnesylation Biotinylation Stearoylation Pyroglutamic acid Carboxylation Phosphorylation Pentoses Hexosamines N-acetylhexosamines Oxidation Glutathionylation Formylation Lipoic acid Myristoylation Palmitoylation Geranylgeranylation Deamidation Sulphation Deoxyhexoses Hexoses Sialic acid

Acids & amides Hydroxyl groups Carbohydrates

Summary
 Post-translational
 Correct

modifications – necessary for protein function
protein folding  Organism development  Cellular Signalling  Motor Proteins  Regulating degradation  …and much more…