L16: Post-translation Modifications Flashcards
Post translational modification (PTM)
Covalent processing events -> change properties of protein by proteolytic cleavage or by addition of modifying group to one or more AA
Change size, charge, structure & conformation of proteins
Involve AA residues
Consequence of PTM
Protein folding/conformation (modification may be important for 3D structure)
Regulation of activity (modification may turn activity on or off)
Protein-protein interaction
Subcellular localisation (modification site may be targeting signal and may be a membrane anchor)
Protein degradation (modification may identify protein for degradation)
Covalent modification of..
N terminus (formulylation, acetylation, pyroglutamate formation)
C terminus (GPI-anchoring, amidation, polyglycylation)
AA residues (side chains)
Peptide bonds
Major types of PTMs
Proteolysis
Phosphorylation
Lipidation
Glycosylation
Proteolytic cleavage
Most common form of PTM
Partial proteolysis of proteins: common maturation step
E.g. insulin. Schematic processing of preproinsulin to proinsulin by signal peptidase in ER
Proinsulin to insulin by proteases in Golgi network
Advantages over synthesis and binding of 2 separate polypeptides
Ensure production of equal amts of A & B chains without coordination of 2 translational activities
Proinsulin folds into 3D structure in which cysteine residues placed for correct disulfide bond formation
Phosphorylation
Most common mechanism of regulating protein function & used for transmitting signals throughout cell
Critical roles in regulation of many cellular processes including cell cycle, growth, apoptosis & signal transduction pathways
Protein kinases
Substrates include lipids, carbohydrates, nucleotides and proteins
Serine, threonine, tyrosine
Phosphoprotein phosphatases
2 major families: PP1 & PP2A
How does phosphate group affect protein function?
Cause conformational changes in phosphorylated protein
- > regulate catalytic activity of protein m
- > recruit neighbouring proteins that recognise and bind to phosphomotifs
Lipidation
Many proteins undergo covalent alterations before they become functional. Conversion of inactive apo forms of proteins by covalent installation
Lipid-anchoring motifs
One or more lipid anchors that help to target the modified proteins to particular membranes
Purpose: anchors proteins to membranes, facilitates protein protein interaction
Types: Palmitoyl group on internal Cys (or Ser), N-Myristoyl group on amino-terminal Gly, Farnesyl (or geranylgeranyl) group on carboxyl-terminal Cys
Exterior: GPI anchor on carboxy terminus
Lipidation-anchoring motifs example
N-myristoylation or amide-linked myristol anchor
Always myristic acid (14 carbon fatty acid)
Always N-terminal
Always a Gly residue that links
Glycosylation
Glycoproteins consist of proteins covalently linked to carbohydrate
Glycosylation classified into 2 groups
O-linked oligosaccharides (O-glycans): O-glycosidic bond; no discernable AA sequence motif
N-linked oligosaccarides (N-glycans): N-glycosidic bond; consensus site or motif on protein is Asn-X-Ser/Thr
N-linked saccharides
High mannose type- contains all mannose (Man) outside core in varying amts
Hybrid type- contains various sugars such as galactose (Gal) and amino sugars such as N-acetyl glucosamine (GlcNAc)
Complex type- is similar to hybrid type. Contains sialic (Sia) acids to varying degrees
Common pentasaccharide core and are synthesised from common precursor oligosaccharide
Synthesis of N-linked saccharides
In ER
Complex carbohydrate chain with 23 or more separate enzyme steps involved in assembly, trimming and maturation of branched carbohydrate structures
Carbohydrate core synthesised -> attached to growing protein
N linked glycoproteins
Functions:
Proper of folding of newly synthesised proteins: antibiotic tunicamycin block N-glycosylation -> non-functional proteins
Proteolytic cleavage example: Insulin
Schematic processing of proinsulin to proinsulin by signal peptidase in ER
Prepoinsulin (insulin synthesised as one long polypeptide) -> cleavage -> proinsulin -> cleavage insulin with c peptide as by product & disulfide bonds form between A & B chain (cysteine forms bond)
Measure c peptide as diabetes test
Phosphorylation example: Glycogen Phosphorylase
Glycogen phosphorylase : an enzyme that breaks down glucose 1 phosphorylate
Glycogen + Pi -> glycogen (n-1) + glucose 1-phosphate
Non-phosphorylated : exist as dimer. Change in conformation for activation. Peptide blocks access to active site -> moves aside -> substrate (phosphate residue) can enter active site
Lipidation example: pyruvate dehydrogenase
Pyruvate -> cleavage -> CO2 -> attach TPP
Disulfide forms on lipoate with redox reaction (acetyl groups transfer to thiol group sitting on lipoate)
Enzyme catalysed reaction: coenzyme A attached to thiol group -> acetyl CoA
Reduce lipoate -> thiol groups reoxidise by dehydrogenase
Lipidation example
Hormone bind to receptor
Hydrophobic lipid group bind to anchor alpha -> GTP binds to alpha -> subunits dissociate from receptor
If alpha wasnt anchored -> drift into cytoplasm
GTP can move from through membrane -> interact with adenyl cyclase (cause synthesis cAMP)
Overtime, GTP breaks down to GDP -> reassociation of complex -> inactivation of G protein and lose hormone
